Method of attaching an antimicrobial cationic polyelectrolyte to the surface of a substrate

ABSTRACT

A method of bonding an antimicrobial cationic polyelectrolyte to the surface of a substrate is described, wherein the antimicrobial thus attached to the substrate provides the substrate with antimicrobial properties, and at least a portion of the bonded antimicrobial is substantially non-leachable during normal conditions of use and storage. A method of manufacturing an antimicrobial material is described which comprises exposure of the substrate to a solution of antimicrobial cationic polyelectrolyte, followed by drying the exposed substrate thoroughly to impart a non-leaching property to at least a portion of the antimicrobial cationic polyelectrolytes.

FIELD OF THE INVENTION

The invention relates to methods of manufacture of inherentlyantimicrobial materials.

BACKGROUND ART

Reduction or elimination of microorganisms on surfaces is important in abroad variety of applications. One approach to interfere with theability of microorganisms to survive on various materials is to modifythe surface of those materials by attachment of antimicrobial agents.

Deciding how best to attach an antimicrobial agent to a material isguided, at least in part, by the planned end-use of the material. Oneimportant and useful consideration is that the antimicrobial activity bepersistent. This may be achieved by permanently attaching theantimicrobial agent to the surface, so that it is unable to migrate orleach away from the modified material surface when the modified materialis exposed to fluids. For example, for applications in which themodified material will come into contact with aqueous fluids, it isimportant that the antimicrobial agent is not rinsed away when themodified material comes into contact with aqueous fluids. Forapplications in which the modified material will come into contact withaqueous biological fluids, it is important that the antimicrobial agentis not rinsed away, or otherwise inactivated, when the modified materialis exposed to aqueous biological fluids. For applications in which themodified material is to be used repeatedly, it is important that theantimicrobial agent is not washed or rinsed away when the modifiedmaterial is washed or rinsed in fluids in between repeated uses. Anadditional consideration in the development of approaches intended tomeet the needs of these and related applications, is that somemicroorganisms have been found to possess the ability to developresistance to certain antimicrobial agents, such as antibiotics orsilver. Currently available approaches do not adequately address all ofthese considerations.

By their design, approaches utilizing leachable active agents (such astriclosan, silver compounds, or biguanides) to impart antimicrobialactivity to materials suffer from eventual depletion or loss of theantimicrobial activity conveyed by the leachable active agents.Depletion or loss of antimicrobial activity can occur especially whenthe materials come into incidental contact with fluids, or duringintentional contact with fluids during washing or rinsing proceduresemployed between repeated uses of the modified material. In addition,the leaching of certain active agents may prohibit or limit the use ofthese approaches in applications where the leaching of the active agentwill cause undesired consequences (e.g. leaching into open wounds,leaching onto products intended for human consumption, and staining ofskin).

One example of an approach utilizing a potentially leachable activeagent is Burba et al. (U.S. Pat. No. 5,154,932), which discloses amethod for providing antimicrobial activity to a formulation or productwhich have negative surface charges, effective for deactivatingmicroorganisms, said method comprising adding to the formulation orproduct an amount of a positively charged layered crystalline mixedmetal hydroxide sufficient to impart antimicrobial activity to theformulation or product.

Another example of an approach utilizing a potentially leachable activeagent is Lyon et al. (U.S. Pat. No. 6,042,877), which discloses a methodof making an antimicrobial article comprising providing a substrate,forming a solution comprising a chelating polymer and a metal ion,depositing the solution on the substrate, drying the substrate to form acoated substrate, and adding a potentiator to the coated substrate toform the antimicrobial article.

Other approaches have employed methods that attach silane-basedquaternary ammonium compounds to particular substrates via a siloxanebond. For example, AEGIS Environments' product line includes productsthat utilize polymers of 3-(trimethoxysilyl)propyldimethyl octadecylammonium chloride. According to product literature, AEM 5700 is 43%3-(trimethoxysilyl) propyldimethyloctadecyl ammonium chloride inmethanol, which can be used to coat the surface of textiles. This is nota polymeric compound; although, some interlinking of the applied silanemay occur after application to the substrate. These types of materialstend to impart a hydrophobic character to the substrates, and are thusless than suitable for many applications. Also, the hydrolyticinstability caused by the bulky (C18 quat) substituent on the siloxanetends to make the materials prone to lose their antimicrobial activity.

Another example utilizing silane-based quaternary ammonium compounds isBlank et al. (U.S. Pat. No. 5,035,892), which discloses a method ofinhibiting the proliferation of potentially destructive microorganismson a substrate comprising treating the substrate with an effectiveamount of an antimicrobial superabsorbent composition formed of acrosslinked hydrophilic sodium salt form of a partially neutralizedacrylic acid based polymer gel, the polymer gel having covalently bondedthereto an organosilane, the organosilane being present in an amount toprevent hydrophobing and reduction of the absorbent capacity of thepolymer gel. Unfortunately, in practice, the amount of organosilane mustoften be reduced to below a desirable antimicrobially efficacious levelin order to prevent the aforementioned hydrophobing. Another example isBlank (U.S. Pat. No. 4,847,088), which discloses a method of inhibitingthe proliferation of potentially destructive microorganisms on asubstrate comprising treating the substrate with an effective amount ofthe synergistic antimicrobial composition comprising a mixture of (a) anorganosilane; and (b) an acid. Again, this example uses siloxanes asdescribed above.

The silane-based quaternary ammonium systems suffer from other drawbacksas well. The long alkyl chains used (typically C-18) tend to make thetreated material hydrophobic—not a desirable property for an absorbentmaterial such as a wound dressing. Additionally, it has been found thatthe silane-quats are deactivated in the presence of blood or otherproteinaceous material (see EP 0136900).

Another example is Shiau et al. (U.S. 010043938 A1), which discloses aprocess for producing an antimicrobial article comprising (a) dissolvinga predetermined amount of quaternary ammonium organosiloxane salt inwater; to make a solution of about 0.05 to 20 wt. % of the salt, (b)mixing a ground calcining aid with a solvent to make a solution, whereinsaid calcining aid to solvent ratio is from 1:1 to 1:10, (c) soaking apreformed honeycomb-shaped substrate into the solution of the aforesaidstep b; followed by drying and calcining at 400-1500° C., (d)impregnating the calcined honeycomb-shaped substrate with the saltsolution of the aforesaid steps a and e, drying the impregnatedsubstrate at 50° to 200° C. These excessive temperatures (400° to 1500°C.) are clearly unsuitable for common substrates such as cellulose andpolymers.

These shortcomings have been overcome, in part, by Batich in U.S. Pat.No. 7,045,673 and application U.S. Ser. No. 020177828 A1. The materialsand methods described in those references teach that an antimicrobialmaterial can be prepared by graft polymerization of an antimicrobialquaternary ammonium monomer onto a substrate such as cellulose. Still,that method has limitations and shortcomings. Namely, it requires avinyl polymerization step which must be conducted under an oxygen-freeatmosphere, which is a costly restriction to commercial development.Furthermore, the process described therein is wasteful in terms ofutilization of antimicrobial cationic polyelectrolyte (for two reasons).First, a large excess of polymerizable quaternary ammonium compound mustbe utilized, and most of that is not incorporated into the finalproduct. Second, the required grafting levels (up to more than 40%antimicrobial “add-on”) are at least an order of magnitude greater thanis required by the present invention, in order to obtain the same levelof antimicrobial efficacy. These shortcomings are discussed furtherelsewhere in the current application.

Lin describes (“Mechanism of Bactericidal and Fungicidal Activities ofTextiles Covalently Modified with Alkylated Polyethyleneimine”Biotechnology and Bioengineering v83(2), p168-172, Jul. 20, 2003) aprocess for covalently attaching hydrophobic polycations to woventextiles using a six-step process that requires various organic solventsand long processing times. The observed microbicidal activity indicateda range of 88% to 99% reduction for various organisms. The processdescribed by the current application is shorter, faster, safer, and moreeconomical than the process described by Lin. In addition, the materialproduced by the method of the current invention is orders of magnitudemore effective in reducing the microbial activity, and is nothydrophobic. The shortcomings of the method described by Lin are citedin the following reference: Daewon Park, Jun Wang, and Alexander M.Klibanov, “One-step, Painting-Like Coating Procedures To Make SurfacesHighly and Permanently Bactericidal” Biotechnol. Prog. 22, p584-589(2006). A similar process was used by Lee (Biomacromolecules 5 p877-883(2004)).

Abel (“Preparation and investigation ofantibacterial carbohydrate-basedsurfaces” Carbohydrate Research 337 (2002) p2495-2499) describes thecovalent attachment of low molecular weight (non-polymeric) quaternaryammonium moieties to cellulose substrates. Like the methods of Lin andLee, Abel's method is a multistep process which requires the use ofhazardous and/or flammable organic reagents and solvents such asp-toluenesulfonylchloride, pyridine, and acetonitrile.

BacStop™ is sold as a fabric sanitizer by Edmar Chemical Company(Cleveland, Ohio). The product contains 50% didecyldimethylammoniumchloride (DDDMAC). It is designed to be added to the final rinse cycleof a laundering process. BacStop™ is claimed to reduce bacterial countby 99.9% (3-log reduction) for Staph. aureus and Klebsiella pneumoniae,and to impart a residual bacteriostatic finish to fabrics. Such aresidual effect is not unexpected, in that several rinses with freshwater would be needed for full removal of just about any solublechemical from a fabric to which it has been applied. The activeingredient (DDDMAC) is a non-polymeric quaternary ammonium compound. TheBacStop™ product label does not claim a residual antimicrobial propertyfor fabrics that have been treated with the product—only a residualbacteriostatic property is claimed. It is demonstrated in comparativeexamples presented below that monomeric quaternary ammonium compoundssuch as DDDMAC are not effective in the practice of the currentinvention, as they do not produce a non-leaching bond with a substrateto give an inherently antimicrobial material with a desirable degree ofantimicrobial efficacy.

Senka, in JP 09078785 describes some of the shortcomings of silane-basedquaternary ammonium antimicrobials for the preparation of inherentlyantimicrobial materials (such as those described by Blank—see above).Senka teaches that an antimicrobial coating can be formed onnon-celluosic substrates using a copolymer of a quaternary ammoniumcompound and acrylic acid derivatives. The copolymer has the propertythat it is soluble in aqueous solution; however, once the solution isdried, the resulting dried solid polymer becomes insoluble. Suchbehavior can be described as “self-crosslinking” in that the polymer (orcopolymer) spontaneously crosslinks upon drying. Thus, a solution of thecopolymer can be applied to a substrate and after drying an insolublecoating with antimicrobial properties is produced. Because the formationof the coating does not depend on interactions between the substrate andthe coating, it is possible to apply this coating to inert substratessuch as synthetic polymers. One skilled in the art would realize thatthese coatings are expected to be flexible when wet, or sufficientlyhydrated; however, they would be expected to be stiff and brittle whendried. This is likely to cause undesirable changes in the physicalproperties of the material, such as stiffness, feel or hand. It is alsolikely that distortion, stretching, or folding of the underlyingsubstrate would likely cause the dry coating to fall apart and detachfrom the substrate, particularly since there is no specific bondingbetween the coating and substrate. The dried copolymer presumably hasbecome crosslinked due to interaction of the positively-chargedquaternary components and the acrylic acid derived components, which areexpected to be negatively-charged at neutral pH. Interaction of theoppositely-charged components of the copolymer is also likely to causesome screening of the positive charge provided by the quaternarycomponent, and resulting in a reduction of antimicrobial efficacy.Furthermore, the simple fact that the coating consists of anothercomponent in addition to the quaternary component necessarily dilutesthe charge density provided by the quaternary component, and thus alsowill reduce the ultimate antimicrobial effect compared to a polymer thatconsists of 100% quaternary component. These shortcomings are discussedfurther below.

In general, coatings are not a desirable approach to modification ofcellulosic substrates such as textiles and wound dressings. While somecoatings may form a very strong bond with a substrate, the attractiveforces responsible for a robust and useful coating are generally betweencomponents of the coating (within the coating itself) rather thanbetween the coating and the substrate. Coatings are generally somewhatthick (like paints, for instance), and can drastically affect thesurface properties of textiles or other substrates. This can happen, forinstance, by blocking or filling-in of porosity, or cementing-togetherof individual fibers. Additionally, some coatings require a curing stepafter application, in order to prevent subsequent dissolution.

Sawan (U.S. Pat. No. 6,264,936) describes an antimicrobial materialwhich can be used to form on the surface of a substrate an antimicrobialcoating or layer which kills microorganisms on contact. Theantimicrobial coating or layer, characterized in the reference as“non-leaching,” is a combination of an organic matrix immobilized on thesurface of the substrate to having biocidal metallic materialsassociated with the matrix. When a microorganism contacts the coating orlayer, the biocidal metallic material is transferred to themicroorganism in amounts sufficient to kill it. Specifically, themetallic antimicrobial agent used is silver. Although this methodpurports to provide a “non-leachable” coating, the mere fact that themetallic antimicrobial agent “is transferred” to the microorganism iscontrary to the common definition of non-leachable. Furthermore, it isknown that although silver and silver salts have very low solubility,the mechanism of antimicrobial activity is dependent on a finitesolution concentration of silver ions. Indeed, Sawan later (column 3,line 9) qualifies the above statement to read “substantially lowleachables”. In a preferred embodiment of Sawan's patent, the organicmaterial comprises a polyhexamethylene biguanide polymer which iscrosslinked with an epoxide, such as N,N-bismethylene diglycidylaniline,to form a crosslinked network or matrix. This crosslinking step isnecessary to prevent dissolution of the matrix. The materials describedby Sawan generally require a curing step, generally in the range of 80°to 120° C., which is unsuitable for many substrates, particularly humanskin. Furthermore, the preferred organic matrix polymer(polyhexamethylene biguanide) is known to be toxic to human cells inhigh concentrations (see U.S. Pat. No. 6,369,289 B1). The use of silveras an antimicrobial agent also incurs some undesirable effects. Onedisadvantage to this approach is that certain bacteria have been able todevelop resistance to silver. (Silver S., “Bacterial silver resistance:molecular biology and uses and misuses of silver compounds.” FEMSMicrobiology Reviews. 2003; 27:341-353). Another disadvantage to thisapproach is that diffusing silver may be able to enter the wound and maypotentially stain the skin. An additional disadvantage of silver is thehigh cost of the raw material. Similar approaches are described in U.S.Pat. Nos. 6,180,584; 6,126,931; 6,030,632; 5,869,073, 5,849,311; and5,817,325.

Brown (EP 0136900) describes a nonwoven fabric with antimicrobialproperties for use as a surgical drape. This is produced by treatment ofa rayon or woodpulp material with a binder and polyhexamethylenebiguanide (PHMB). Because the applied PHMB is extractable by aqueousfluids, the amount of PHMB must be kept below a critical level, in orderto prevent the leached PHMB from reaching toxic levels.

Payne, in U.S. Pat. No. 5,700,742, describes treatment of textilematerials with combinations of PHMB and a strong acid in order toovercome problems such as discoloration, loss of antimicrobial efficacy,and undesirable changes to substrate properties that are associated withtreatment of textiles using PHMB without the added strong acid.

Orr (U.S. Pat. No. 6,369,289 B1) teaches the use of PHMB in cellulosicwound dressings. Orr teaches that leachable PHMB can lead to adverseeffects such as skin disorders, redness, tenderness and hives, but thatthese are avoided by utilizing a precisely-controlled amount to PHMB inthe wound dressing. Non-leaching of the applied antimicrobial is notdemonstrated or even suggested. Orr merely teaches that the amount ofleachable PHMB is less than that which would cause irritation to skin oran open wound; however he provides no data to this effect, or that thematerials provide useful antimicrobial efficacy at the level of PHMBused. In fact, Orr calculates the amount of PHMB applied to the dressing“by extraction”, which necessarily implies that the materials can beleached or extracted. The use levels of PHMB cited by Orr are onlyslightly below those claimed by Brown (EP 0136900).

The method of Orr (U.S. Pat. No. 6,369,289 B1) is used to produce acommercial antimicrobial wound dressing known as “Kerlix-AMD”, whichcontains 0.2% PHMB. The Kerlix-AMD dressing is known to show a distinctzone-of-inhibition (ZOI) in a Kirby-Bauer test (see Kerlix-AMD productbrochure available at(http://www.kendallhq.com/catalog/brochures/KerlixSS.pdf). A measurableZOI is a definite indicator of leachable antimicrobial activity. Theantimicrobial effectiveness of Kerlix-AMD has been described“Effectiveness of a New Antimicrobial Gauze Dressing as a BacterialBarrier” A. M. Reitsma, et al., University of Virginia Health System,Charlottesville, Va. This study makes no mention of non-leachableproperties.

PHMB has been studied as an antimicrobial treatment for cotton fabric(“Testing the Efficacy of polyhexamethlylene Biguanide as anAntimicrobial Treatment for Cotton Fabric,” Michelle Wallace, AATCCReview p18-20, November, 2001). Antimicrobial efficacy was maintainedafter laundering; however, no discussion of leaching is given; however,the data does show that antimicrobial efficacy diminishes with repeatedlaundering cycles. Presumably, this is due to loss of PHMB from thesubstrate (leaching).

Consequently, there exists a need for a method that can attach aneffective and non-leaching amount of antimicrobial agent to a variety ofsubstrate materials in a convenient, reliable, and cost-effectivemanner. The inadequacies of existing approaches are overcome with thepresent inventive method wherein an improved method of non-leachablyattaching antimicrobial agents to a variety of substrate materials isprovided.

SUMMARY OF THE INVENTION Industrial Applicability

It is an aspect of this invention to provide a method of manufacturingan inherently antimicrobial absorbent material. The method establishesthe attachment of non-leaching antimicrobial cationic polyelectrolytesto substrates, thereby imparting to the substrates an inherentnon-leaching antimicrobial property, and not causing unwantedside-effects such as making the material hydrophobic (water repellent).The method of manufacturing an inherently antimicrobial materialincludes exposing the substrate to an aqueous solution of antimicrobialcationic polyelectrolytes, followed by drying the exposed substratethoroughly to impart a non-leaching property to at least a portion ofthe antimicrobial cationic polyelectrolytes.

It is an aspect of the current invention to provide a method ofmanufacturing an inherently antimicrobial material comprised of asufficient number of polymeric diallyldimethylammonium chloridemolecules, non-leachably attached to a substrate comprised in whole orin part of a celluosic material, to render the material antimicrobialand non-hydrophobic, before, during, and after exposure of the materialto aqueous fluids, said method comprising; loading the substrate withsaid polymeric diallyldimethylammonium chloride molecules by exposingthe substrate to an aqueous solution of said polymericdiallyldimethylammonium chloride molecules, and thoroughly drying theloaded substrate to impart a non-leaching property to at least a portionof said polymeric diallyldimethylammonium chloride molecules.

It is an aspect of the current invention to provide a method ofmanufacturing an inherently antimicrobial material comprised of asufficient number of antimicrobial cationic polyelectrolytes,non-leachably attached to a substrate, to render the materialantimicrobial and nonhydrophobic, before, during, and after exposure ofthe material to aqueous fluids, said method comprising; loading thesubstrate with said antimicrobial cationic polyelectrolytes by exposingthe substrate to an aqueous solution of said antimicrobial cationicpolyelectrolytes, and thoroughly drying the loaded substrate to impart anon-leaching property to at least a portion of said antimicrobialcationic polyelectrolytes.

It is an aspect of this invention to provide a method of attaching anantimicrobial cationic polyelectrolyte to a substrate wherein saidsubstrate is comprised in whole or in part of cellulose, and whereinsaid antimicrobial cationic polyelectrolyte is not self-crosslinking andhas an average degree of polymerization of at least 3, wherein saidmethod comprises the steps of wetting the substrate with an aqueoussolution of the antimicrobial cationic polyelectrolyte followed bythoroughly drying of the wetted substrate, wherein said drying causes atleast a portion of the antimicrobial cationic polyelectrolyte to becomeattached to the substrate in a non-leachable manner, and wherein theattached antimicrobial cationic polyelectrolyte provides anantimicrobial effect to the resulting product.

It is an aspect of this invention to provide a method of impartingnon-leachable antimicrobial properties to a substrate wherein saidsubstrate is comprised in whole or in part of cellulose, and whereinsaid non-leachable antimicrobial properties result from strongattractive interactions between said substrate and an appliedantimicrobial cationic polyelectrolyte wherein said strong attractiveinteractions are caused to occur by thoroughly drying of the substrateafter application of an aqueous solution of said antimicrobial cationicpolyelectrolyte to the substrate.

It is an aspect of the current inventive method to provide a method ofmanufacturing an inherently antimicrobial absorbent material.

It is an aspect of this invention that thorough drying is accomplishedby application of infrared heat, radiant heat, or hot air.

It is an aspect of the current inventive method that an additional stepof rinsing, washing, or extracting will remove any leachable unbondedportion of said antimicrobial cationic polyelectrolyte from theinherently antimicrobial material.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are polymeric molecules containing at leastthree quaternary ammonium active units per molecule.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are polymeric molecules having an averagedegree of polymerization selected from the group consisting of 3 to25,000, 20 to 10,000, and 100 to 2500.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are defined as polymeric molecules having amultiplicity (i.e. more than three) of cationic charges per polymericmolecule, or a net cationic charge excess of greater than three chargesper polymeric molecule. “Net cationic charge excess” is defined as thesum of all the cationic charges in a given molecule minus the sum of allnegative charges in the same molecule, not including the charges of anyassociated counterions (such as chloride ions) which are not covalentlybonded to the polymeric molecule, and shall be considered to be apositive number equal to or greater than three, for the purposes of thisinvention.

Charge density is a measure of the relative amount of cationic charge ina given cationic polyelectrolyte, and higher antimicrobial efficacy willgenerally correlate with higher charge density for a given polymer. Itis an aspect of this invention that the cationic polyelectrolytes have aminimum excess cationic charge of greater than approximately 1 mole (1mole equals 6.02×10²³) per 25,000 grams of cationic polyelectrolyte (theweight of cationic polyelectrolyte includes the weight of the polymercomponent plus the weight of any associated counterions). Preferably,the cationic polyelectrolytes have a minimum excess cationic charge ofgreater than approximately 1 mole per 2,500 grams of cationicpolyelectrolyte. More preferably, the cationic polyelectrolytes have aminimum excess cationic charge of greater than approximately 1 mole per500 grams of cationic polyelectrolyte. Even more preferably, thecationic polyelectrolytes have a minimum excess cationic charge ofgreater than approximately 1 mole per 212 grams of cationicpolyelectrolyte. Most preferably, the cationic polyelectrolytes have aminimum excess cationic charge of equal to greater than approximately 1mole per 162 grams of cationic polyelectrolyte.

Zeta potential refers to the electrostatic potential generated by theaccumulation of ions at the surface of a colloidal particle which isorganized into an electrical double-layer consisting of the Stern layerand the diffuse layer of a material. Zeta potential and instruments usedto measure zeta potential are well known in the art. The zeta potentialof a successfully treated substrate should be significantly higher thanthat of an untreated substrate, due to the presence of the antimicrobialcationic polyelectrolyte. Thus, the antimicrobial efficacy of a treatedsubstrate can be determined by measuring its zeta potential. It is anaspect of the current inventive method that the zeta potential of theinherently antimicrobial material produced is negative (less than zero).

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are polymeric phosphonium compounds containingat least three quaternary phosphonium active units per molecule.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are comprised, in whole or in part, ofmonomeric units having the structure CH₂═CR—(C═O)—X—(CH₂)n^(−N)⁺R′R″R′″//Y⁻; wherein, R is hydrogen or methyl, n equals 2 or 3, X iseither O, S, or NH, and R′, R″ and R′″ are independently selected fromthe group consisting of H, C1 to C16 alkyl, aryl, arylamine, alkaryl,and aralkyl, and Y⁻is an anionic counterion to the positive charge ofthe quaternary nitrogen; diallyldimethylammonium salts; vinyl pyridineand salts thereof; vinylbenzyltrimethylammonium salts;diallyldialkylammonium salts and vinylbenzyltrialkylammonium salts.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are comprised, in whole or in part, ofmonomeric units having the structure CH₂═CR—(C═O)—X—(CH₂)_(n−)NR′R′″;wherein, R is hydrogen or methyl, n equals 2 or 3, X is either O, S, orNH, and R′ and R″ are independently selected from the group consistingof H, C1 to C16 alkyl, aryl, arylamine, alkaryl, and aralkyl.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are polymeric molecules known as polyDADMACcomprised, in whole or in part, of diallyldimethylammonium chloride,(also known as DADMAC).

It is an aspect of this invention that said antimicrobial cationicpolyelectrolytes are comprised of polyDADMAC homopolymer or polyVBTAChomopolymer, where “polyVBTAC” means poly(vinylbenzyltrimethylammoniumchloride). “Homopolymer” is defined as a polymeric material consistingof multiple units derived from a single type of polymerizable monomer.

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are polymeric molecules comprised, in whole orin part, of monomeric units selected from the group consisting ofdimethylaminoethyl methacrylate, dimethylaminoethyl methacrylate(hydrogen chloride quaternary), dimethylaminoethyl methacrylate (methylchloride quaternary) and dimethylaminoethyl methacrylate (benzylchloride quaternary).

It is an aspect of the current inventive method that said antimicrobialcationic polyelectrolytes are comprised, in whole or in part, ofmonomeric units selected from the group consisting of dimethylaminoethylacrylate, dimethylaminoethyl acrylate (hydrogen chloride quaternary),dimethylaminoethyl acrylate (methyl chloride quaternary) anddimethylaminoethyl acrylate (benzyl chloride quaternary).

It is an aspect of the current inventive method that said substrate iscomprised, in whole or in part, of cellulose.

It is an aspect of the current inventive method that said substrate iscomprised, in whole or in part, of at least one cellulosic materialselected from the group consisting of hydroxyethyl cellulose,carboxymethyl cellulose, methyl cellulose, rayon, cotton, linen and woodpulp.

It is an aspect of the current inventive method that said inherentlyantimicrobial material has an absorbent capacity for aqueous fluids.

It is an aspect of the current inventive method that said inherentlyantimicrobially material has an absorbent capacity for aqueousbiological fluids.

It is an aspect of the current inventive method that said substrate is awoven, flexible material.

It is an aspect of the current inventive method that said substrate is anon-woven, flexible material.

It is an aspect of the current inventive method that said substrate iswood or lumber.

It is an aspect of the current inventive method that said substrate ispaper.

INDUSTRIAL APPLICABILITY

It is an aspect of the current inventive method that said inherentlyantimicrobial material comprises all or part of a wound dressing, a burndressing, a sanitary pad, a tampon, an intrinsically antimicrobialabsorbent dressing, a diaper, toilet paper, a sanitary wipe, a sponge, acotton swab, a surgical gown, an isolation gown, a lab coat, a glove,surgical scrubs, a head cover, a hair cover, a face mask, a suture, afloor mat, a lamp handle cover, an exam table cover, a cast liner, asplint liner, padding, gauze, blood transfer tubing, a blood transferstorage container, sterile packaging, a mattress cover, bedding, asheet, a towel, clothing, underwear, a sock, shoe-cover, an automobileair filter, an airplane air filter, an HVAC system air filter, amilitary protective garment, an apparatus for protection against abiohazard or biological warfare agent, lumber, food packaging material,meat packaging material, fish packaging material, apparel for foodhandling, a surface for food preparation, a contact lens, carpet, wood,lumber, paper, or paper currency.

Definitions

“Microbe” or “microorganism” refers to any organism or combination oforganisms such as bacteria, viruses, protozoa, yeasts, fungi, molds, orspores formed by any of these.

“Antimicrobial” refers to the microbicidal or microbistatic propertiesof a compound, composition, article, or material that enables it tokill, destroy, inactivate, or neutralize a microorganism; or to preventor reduce the growth, ability to survive, or propagation of amicroorganism.

“Substrate” refers a surface or medium upon which an antimicrobialpolyelectrolyte is chemically bonded.

“Cationic polyelectrolyte” means a polymer molecule with multiplecationic sites or moieties which are covalently bonded to the polymer,or attached to the molecular structure of the antimicrobial polymer bycovalent chemical bonds, and are part of the polymer molecularstructure, and that said cationic sites or moieties are located eitherin the main-chain of the polymer, or in side-groups of the polymer.“Main-chain” and “side-groups” are terms commonly used to describepolymer molecular structure and will be familiar to one skilled in theart.

The term “quaternary ammonium” is common chemical nomenclature and itmeaning will be understood by one skilled in the art. There are twotypes of quaternary ammonium compounds: acidic, and non-acidic. Acidicquaternary ammonium compounds are acid salts of amines, and arecharacterized by having a N—H covalent bond wherein the N—H bond isreactive with bases. Non-acidic quaternary ammonium compounds do nothave this N—H bond, and are not reactive with bases. Non-acidicquaternary ammonium compounds are preferred in the practice of thisinvention.

By “inherently antimicrobial” is meant a property of a material whereinsaid material would exhibit antimicrobial activity or properties in theabsence of any antimicrobial activity or properties contributed byagents, compounds, or additives which are not integral to the material,not chemically bonded to the material, or detachable from the material,or after the removal or depletion of such agents, compounds, oradditives from the material. “Inherently antimicrobial” does not meanthat the material contains no leachable agents with antimicrobialactivity.

By “non-leaching” is meant that the antimicrobial cationicpolyelectrolytes of the present invention, once attached to the materialor substrate via the method of the current invention, do not appreciablyseparate from, migrate out of, or away from the material or substrate,enter a wound, or otherwise become non-integral with the material orsubstrate under standard uses. By “not appreciably separate” is meantthat no more than an insubstantial amount of antimicrobial cationicpolyelectrolyte separates, for example less than one percent, preferablyless than 0.1 percent, more preferably less than 0.01 percent, and evenmore preferably less than 0.001 percent of the total quantity ofantimicrobial cationic polyelectrolyte. Alternatively, “not appreciablyseparate” means that the solution concentration of antimicrobialcationic polyelectrolyte resulting from separation of attachedantimicrobial cationic polyelectrolyte, in a liquid in contact with thematerial or substrate, does not exceed a predetermined level, forexample less than 0.01%, preferably less than 0.005%, and morepreferably less than 0.001%. Alternately, depending on the application,“not appreciably separate” may mean that no adverse effect on woundhealing or the health of an adjacent tissue of interest is measurable.It should be understood that particular definition may depend on theapplication in which the invention is used. For instance, in textileapplications, the desire is to maintain efficacy over a prolonged periodof use, thus only a very gradual loss of antimicrobial material over anextended time would be acceptable, regardless of the amount leached atany given point in time. For medical applications such as wounddressings, the overriding concern would be to ensure that the localizedconcentration of leachable material remains below a specific level at agiven point in time, or leads to no adverse effects over the period ofuse.

In regard to the foregoing definition, it is noted that “non-leachable”refers to the bond between the polymer chain and the substrate. Incertain embodiments of the present invention, a bond between the polymerbackbone and one or more type of antimicrobial group may beintentionally made to be more susceptible to release, and therefore moreleachable. This may provide a benefit where it is desirable for apercentage of the antimicrobial groups to be selectively released undercertain conditions. However, it is noted that the typical bond betweenthe polymer chain and antimicrobial groups envisioned and enabled hereinare covalent bonds that do not leach under standard exposure conditions.

By “degree of polymerization” is meant the number of monomers that arejoined in a single polymer chain. For example, in a preferred embodimentof the invention, the average degree of polymerization is in the rangeof about 5 to 1,000. In another embodiment, the preferred average degreeof polymerization is in the range of about 10 to 500, and in yet anotherembodiment, the preferred average degree of polymerization is in therange of about 10 to 100.

“Self-crosslinking” means that the polymer has the capability to undergoa chemical or physical reaction with itself which results in bridging,bonding, or attachment between its individual polymer chain molecules toform a three dimensional network structure consisting essentially of asingle large molecule, without the need to react with any outsidereagents such as catalysts or crosslinking agents which are not alreadypart of the polymer's molecular structure.

A “cellulosic material” means a natural material made in whole or inpart of cellulose or a synthetic material derived from cellulose orhaving chemical and physical properties similar to cellulose.

DETAILED DESCRIPTION

The current invention provides a method of manufacture of an inherentlyantimicrobial material that establishes a non-leaching attachment ofantimicrobial cationic polyelectrolytes onto a substrate. Thenon-leaching attachment of antimicrobial cationic polyelectrolytes to asubstrate that has been exposed to a solution of antimicrobial cationicpolyelectrolytes (the treatment solution) occurs during the thoroughdrying of the substrate. In one exemplary embodiment of the inventivemethod, a substrate containing cotton, or a cellulose derivative, isthoroughly dried after it has been exposed to an aqueous solution ofpolymeric diallyldimethylammonium chloride (i.e. polyDADMAC) having anaverage degree of polymerization of ranging from 5 to 10,000, or morepreferably 30 to 5,000, and most preferably 100 to 2500.

As used herein, “antimicrobial” refers to the property of a compound,composition, article, or material that enables it to destroy, neutralizeor kill a microorganism. As used herein, “microbe” or “microorganism”refers to any organism or combination of organisms able to causeinfection, such as bacteria, viruses, protozoa, yeasts, fungi, or molds.

It is an aspect of the inventive method that the antimicrobial cationicpolyelectrolytes comprise polymeric phosphonium compounds. Polymericphosphonium compounds are known to possess antimicrobial properties.Several reports in the chemical literature concern the synthesis ofvarious antimicrobial synthetic polymers. For example, the synthesis ofpolymeric phosphonium derivatives of styrene has been reported by Endo,T., et al in “Novel Polycationic Biocides: Synthesis and AntibacterialActivity of Polymeric Phosphonium Salts” (Journal of Polymer SciencePart A: Polymer Chemistry, 31, pp. 335-342, 1993). Phosphoniumquaternary polymers have been shown to be up to 4 orders of magnitudemore effective as antimicrobial agents than the corresponding nitrogenquaternary polymers.

It is an aspect of the inventive method that suitable concentrations ofthe antimicrobial cationic polyelectrolytes in the treatment solutionrange from about 20 wt % to about 0.01 wt %, and preferably between 10wt % and 0.1 wt %. The actual concentration chosen for a particularapplication depends on, among other things, the molecular weight of theparticular antimicrobial cationic polyelectrolytes and the resultantviscosity of the solution. At a minimum, a sufficient amount ofantimicrobial must be incorporated into the product in order to providethe desired degree of antimicrobial activity (as described below). Thereis no distinct upper limit to the concentration of the solution of theantimicrobial cationic polyelectrolytes that can be used, other thanpractical constraints, such as viscosity, solubility, and cost, whichwill vary according to the antimicrobial cationic polyelectrolyteutilized.

It is an aspect of the inventive method that there is no minimumincubation time after the loading of substrate with antimicrobial byexposure of substrate to a solution of antimicrobial cationicpolyelectrolytes. The loading of substrate with antimicrobial cationicpolyelectrolytes is complete as soon as the substrate is wetted withantimicrobial cationic polyelectrolyte solution. Uniformity of wettingallows the final product to have an even and uniform distribution ofattached antimicrobial cationic polyelectrolytes, but is not a necessaryprerequisite for attachment of antimicrobial cationic polyelectrolytesto any specific area of the substrate.

To establish a non-leaching attachment of antimicrobial cationicpolyelectrolytes to a substrate, the substrate which has been loadedwith antimicrobial cationic polyelectrolyte solution must be thenthoroughly dried. Thorough drying is necessary to impart thenon-leaching property conveyed by the inventive method to at least aportion of the antimicrobial cationic polyelectrolytes that were in thesolution contacting the substrate. It is an aspect of the inventivemethod to use any temperature and time combination that results inthorough drying of said material. As used herein, thoroughly driedmeans, for instance, that a substrate exposed to a solution ofantimicrobial cationic polyelectrolytes is then dried to a constantweight. As used herein, dried to a constant weight means dried to thepoint at which continued application of the chosen drying procedure willno longer result in a considerable additional measurable loss of weightdue to evaporation of water or other solvent. Attainment of constantweight is a useful tool to measure extent of dryness; however, theattainment of constant weight is not the actual factor that enablesnon-leachable attachment of the antimicrobial to the substrate. Thatattachment is produced by the drying process itself (removal of waterfrom the system). The particular temperatures and drying times necessaryto achieve thorough drying depend, among other things, on the particularsubstrate material, the initial amount of moisture in the article, theweight and size of the article, the amount of airflow provided to thearticle during drying, and the humidity of the air or other mediumcontacting the article. Illustrative examples are provided below whichdescribe the effect of achieving various degrees of dryness by allowingthe treated substrate to reach moisture equilibrium at specific pointsof relative humidity. Any drying apparatus, drying method, andtemperature and drying time combination that thoroughly dries thetreated substrate and imparts a non-leachable bond between the substrateand antimicrobial is sufficient. For purposes of illustration, dependingon the particular characteristics of a particular application, thedrying step may be performed in an oven (e.g. 80° C. for 2 hours), in ahigh throughput furnace (e.g. 140° C. for 30 seconds), in a clothesdryer, in a desiccator, in a vacuum chamber, in a dehumidifier, in adehydrator, or in a lyophilizer (freeze dryer). Infrared heat, radiantheat, microwave, and hot air are all suitable drying methods for thesubstrate which has been exposed to a solution of antimicrobial cationicpolyelectrolytes. The upper limit of drying temperature for a particularapplication will generally be determined by the degradation temperatureof the particular substrate or antimicrobial cationic polyelectrolytebeing treated.

Unconventional or nontraditional drying methods may be utilized. Forinstance, simply freezing of the wet substrate which has been treatedwith antimicrobial solution has been found to impart a non-leachingbonding of the antimicrobial to the substrate. Presumably, this is dueto the fact that upon freezing the dissolved antimicrobial is pushed outfrom the crystallizing ice structure, leaving the previously dissolvedantimicrobial deposited onto the surface of the substrate, and forming anon-leaching bond between the substrate and antimicrobial cationicpolyelectrolytes. This results in the same overall effect asconventional drying, wherein the water is separated from the polymer byevaporation. As expected, rapid freezing, such quenching the wetmaterial in liquid nitrogen, is not as effective at promotingnonleachable attachment because of insufficient time for to achieveseparation of antimicrobial from water.

A similar effect is observed when the substrate which has been treatedwith an aqueous solution of antimicrobial is subsequently washed with anorganic solvent that is miscible with water, in order to remove water,followed then by washing in water. The conventional drying step may beomitted; however, non-leachable bonding of the antimicrobial to thesubstrate may still be achieved because the solvent removes water in amanner similar to what happens during normal drying. Generally, a watersoluble polymer such as polyDADMAC will be insoluble or less soluble inan organic solvent, compared to water, and the bonding effect will bemore pronounced as the solubility of the antimicrobial polymer in thechosen solvent is lower. For instance, it has been found that t-butylalcohol and tetrahydrofuran promote non-leachable bonding ofantimicrobial polymer to a cotton substrate better than dimethyformamideor methanol.

It is an aspect of this invention that the antimicrobial polymer isapplied to the substrate as an aqueous solution, and that organicsolvents are not required. The use of aqueous solutions is an advantageover the use of organic solvents because of various issues includingcost, safety, health, and regulatory. It is also possible to utilizemixed solvents, such as water/alcohol mixtures, for initial applicationof the antimicrobial to the substrate using the described process,combined with any of the drying methods described above. This willdepend on the solubility of the antimicrobial in the mixed solventsystems. For instance, mixtures of alcohol and water may be used. It mayalso be possible to use completely non aqueous solvent systems; however,it is necessary that the antimicrobial be soluble in the chosen solventsystem.

Other drying methods such as supercritical fluid drying may also besuccessfully employed in the practice of this invention. Freeze dryingmay be used; however, is unnecessary since merely freezing of thesubstrate which has been wetted with antimicrobial is sufficient.Subsequent sublimation or removal of the ice phase is not necessarilyrequired in order to effect a non-leachable bond between the substrateand the antimicrobial cationic polyelectrolytes.

It is an aspect of the inventive method that prior to the drying step,in order to lessen the time needed to thoroughly dry the loadedsubstrate, and/or to reduce consumption of materials, a mechanicalaction or force may be applied to the loaded substrate to remove excessantimicrobial-containing solution from the loaded substrate. Anymechanical action or force may be applied; however, it is preferred thatsuch action or force be uniform in order to provide an even distributionof remaining solution within the loaded substrate as the solution isforced out. Examples of such mechanical action or force include, but arenot limited to, rolling, pressing, squeezing, centrifugation, and thelike. It should be noted application of a mechanical force to removeexcess solution prior to drying is distinct from the drying procedure inthat the mechanical force removes both the antimicrobial and the carriersolution, while the drying procedure removes only the carrier solution,through evaporation, but leaves the antimicrobial in the loadedsubstrate.

The non-leaching attachment of antimicrobial cationic polyelectrolytesproduced by said inventive method has been demonstrated by boiling thetreated articles of this invention in water containing a range of saltconcentrations, and in water having a range or various neutral, acidic,and alkaline pH values, followed by verification that the treatedarticles retain antimicrobial activity. In addition, antimicrobialefficacy of the materials having non-leaching attachment of theantimicrobial cationic polyelectrolyte has been demonstrated afterexposure of the inherently antimicrobial materials to proteinaceousmaterial in the form of fetal bovine serum.

It is an aspect of the current inventive method that a rinsing step maybe optionally exercised. It is likely that when utilizing the method ofthe current invention, that only a portion of the total antimicrobialcationic polyelectrolytes applied to the substrate will actually becomenon-leachably bonded to the substrate; hence, the inherentlyantimicrobial material is likely to also contain some leachableantimicrobial cationic polyelectrolytes. The decision of whether or notto rinse the treated material will depend on whether a leachingantimicrobial property, in addition to the inherently non-leachingantimicrobial property, is desired in the final product. For someapplications (e.g. textile applications), it may be desirable to retainsome or all of the leachable portion of antimicrobial cationicpolyelectrolytes in the final product, in combination with thenon-leaching portion, because the leachable portion can contribute tothe overall antimicrobial activity, at least initially, before theleachable portion becomes depleted. So, for example, where a particularapplication calls for retention of the leachable portion, it is suitableto utilize the inherently antimicrobial material after it has beenthoroughly dried (without rinsing). For other applications (e.g. somewound dressings), it may be desirable to remove the leachable portionand retain only the non-leaching portion. For example, where aparticular application calls for removal of the leaching portion, thethoroughly dried treated material can be repeatedly rinsed in fluid toremove the leachable portion of antimicrobial cationic polyelectrolytesthat did not attach to the substrate during the thorough drying step. Inone exemplary embodiment, the rinsing step can be considered completewhen conductivity readings of the rinsate equal that of the input rinsefluid, indicating that the rinsate is free of unbounds. In anotherexemplary embodiment, the rinsing can be accomplished by using a saltsolution, followed by rinsing in fresh water to remove both theleachable antimicrobial and salt in order to obtain the lowest possiblelevel of leachable antimicrobial.

It is an aspect of the current inventive method that it can establish anon-leaching attachment of antimicrobial cationic polyelectrolytes to avariety of substrates. Natural and synthetic substrate materialsamenable to the current inventive method include, but are not limitedto, cellulose, cellulose derivatives, hydroxyethyl cellulose,carboxymethyl cellulose, methyl cellulose, rayon, cotton, wood pulp,linen, polysaccharide, protein, wool, collagen, gelatin, chitin,chitosan, alginate, starch, silk, polyolefin, polyamide, fluoropolymer,polyvinyl chloride (PVC), vinyl, rubber, polylactide, polyglycolide,acrylic, polystyrene, polyethylene, polypropylene, nylon, polyester,polyurethane, and silicone, all of which may be verified by routineexperimentation based on the present disclosure.

It is an aspect of the current inventive method that the antimicrobialactivity exhibited by materials manufactured by the method is veryrobust. In contrast, some competing formulations, such as those marketedby AEGIS Environments, have been found to be inactivated after exposureto blood (see EP 0136900) or 10% fetal bovine serum. In one exemplaryembodiment of the current inventive method, the antimicrobial activityof non-leachably attached polymeric molecules of the quaternary ammoniumcompound diallyldimethylammonium chloride remains robust in the presenceof 10% fetal bovine serum, as described in the examples below. Thisaspect of the current inventive method will permit antimicrobialactivity to persist in the presence of bodily fluids, which is avaluable and useful property for many applications in the healthindustry.

It is an aspect of this invention that silane, silicone, or siloxaneantimicrobial cationic polyelectrolytes are not applied to the substrateor incorporated into the antimicrobial material, as silane, silicone orsiloxane compounds generally will impart a water-repellent character toa substrate of composition, thus reducing the absorbency of thematerial.

It is an aspect of this invention that the process does not require theuse of an inert atmosphere, vacuum, high pressure, irradiation, organicsolvents, catalysts, excessively high temperatures, and/or volatile,expensive, flammable, or toxic reagents to produce the antimicrobialmaterial. This is in contrast to prior methods which require suchmeasures in order to produce a cellulosic substrate with attachedquaternary ammonium.

It is an aspect of specific embodiments of the current inventive methodthat the attachment of antimicrobial cationic polyelectrolytes to afabric substrate has a softening effect on fabric, and can therebyreduce the amount of softener that must to be applied to the fabric.

It is an aspect of the specific embodiments of the current inventivemethod that cationic fabric softening agents can be added to saidtreatment solution to impart an enhanced antimicrobial effect to theproduct of said inventive method. We have determined that cationicsoftening agents alone do not show any permanent antimicrobial effect,and therefore we regard this observation of the synergic interactionbetween the cationic softening agents and the cationic antimicrobialcationic polyelectrolyte to be a novel and enhancing effect.

Some substrates are more amenable to treatment by selected antimicrobialcationic polyelectrolytes than other similar substrates. For instance,rayon and cotton are both forms of cellulose; however, it has been found(unexpectedly) that cotton treated using polyDADMAC in accordance withthe methods described herein gives a product with a significantly higherantimicrobial power than rayon treated in an identical manner. This isan unexpected result, and it has been found to be particularly useful,for instance, in the manufacture of an antimicrobial wound dressing.Wood pulp is also significantly more reactive than rayon when used inconjunction with polyDADMAC. These differences may not exist, or even bereversed when different antimicrobial cationic polyelectrolytes areemployed.

The strong non-leachable binding of the antimicrobial cationicpolyelectrolytes to the substrates, as described herein, is unexpectedbased on comparisons to other model systems such as formation ofpolyelectrolyte complexes between cationic polymers and clay or micaparticles. For instance, treatment of mica with an aqueous solution ofpolyDADMAC, followed by drying and washing in distilled water results ina material capable of binding an anionic dye, as described in thefollowing examples; however, if this treated material is subsequentlyrinsed in saline solution, the ability to bind anionic dye issubstantially diminished, or completely lost, indicating a loss ofpolyDADMAC from the material. As such, the clay-PolyDADMAC system doesnot constitute an antimicrobial bonded to a substrate in a non-leachablemanner. Thus, the mechanism of binding of the cationic antimicrobial tothe surface as described for the current invention, is presumably notstrictly an ion-exchange type of process, as exhibited by thepolyDADMAC-mica system. Although it is known that cellulose normallyexhibits a net negative surface charge (or zeta potential), it is notconsidered to be an ionic material. Thus, ionic interactions betweencellulose and a cationic species would not be expected to beexceptionally strong or irreversible. Presumably, the stronginteractions which result in non-leachable bonding observed betweencotton and polyDADMAC, for example, are based on a balanced combinationof hydrophobic and ionic interactions. These interactions are formedduring drying or removal of water from the system. The overall strengthof these interactions is compounded by the polymeric nature of thesubstrate and the antimicrobial cationic polyelectrolyte, which resultsin a multiplicity of binding effects. This multiplicity of bindingeffects means that each polymeric antimicrobial molecule is held to thesubstrate through bonding attractions emanating from many differentpoints along the molecular structure, which combine to give a verystrong overall bond between the substrate and the polymericantimicrobial cationic polyelectrolyte. This multiplicity of bindingeffects cannot be achieved for smaller molecules, such as monoquats(benzalkonium chloride, or DDDAMC, for instance). It cannot be ruled-outthat the drying process induces the formation of covalent chemical bondsbetween the substrate and the antimicrobial cationic polyelectrolyte.This could occur, for instance, by condensation (dehydration) reactionsof the hydroxyl groups on cellulose with reactive sites on theantimicrobial molecules. Insertion of portions of the antimicrobialcationic polyelectrolytes into the crystalline structure of thecellulose substrate could also be partially responsible for the observedbinding. Polymeric quaternary ammonium compounds exhibit thenon-leaching attachment described herein. The use of non-polymericquaternary ammonium compounds according to the method of this inventiondoes not produce a nonleachable attachment. Comparative examples of thelack of non-leachable attachment of nonpolymeric quats are given below.In light of the behavior of the nonpolymeric quats, it would not beobvious to one skilled in the art that polymeric quats would becomenon-leachably attached when used in the method of the current invention.

The discovery that drying of a cellulose substrate to which a cationicpolyelectrolyte has been applied causes irreversible, non-leachablebonding between at least a portion of the applied cationicpolyelectrolyte has not previously been described. Onabe (“Studies ofInterfacial Properties of Polyelectrolyte-Cellulose Systems. I.Formation and Structure of Adsorbed layers of CatioinicPolyelectrolyte-(Poly-DMDAAC) on Cellulose Fibers” Journal of AppliedPolymer Science. Vol. 22, 3495-3510 (1978)) studied the adsorption ofpolyDADMAC onto cellulose fibers. The cellulose fibers were soaked inpolyDADMAC solutions for 24 hours, and then rinsed to remove excesspolyDADMAC prior to analysis. Based on the studies and data presentedherein to describe the current invention, it will be recognized thatsuch a rinsing step (without first drying the exposed substrate) doesnot produce an inherently antimicrobial material with cationicpolyelectrolyte non-leachably attached to the cellulose substrate.Apparently, the rinsing procedure described by Onabe is not sufficientto remove all of the applied cationic polyelectrolyte; however, theamount remaining is insufficient to provide antimicrobial efficacy. Thishas been verified, as described in the examples below. Onabe does notdiscuss antimicrobial properties of cellulose materials with adsorbedcationic polyelectrolytes, nor does the reference discuss whether thosematerials show any non-leachable bonding. Onabe attributes absorption ofcationic polyelectrolyte to cellulose as strictly an electrostaticinteraction, which is inconsistent with the observations of thenon-leachable properties of the current invention when exposed to salinesolutions.

It is known; however, that enhanced antimicrobial properties areexhibited by polymeric quaternary compounds, relative to monomeric quats(keda, T., “Antibacterial Activity of Polycationic Biocides”, Chapter42, page 743 in: High Performance Biomaterials, M. Szycher, ed.,Technomic, Lancaster, Pa. (1991); and Ikeda T, Yamaguchi H, and Tazuke,S “New Polymeric Biocides: Synthesis and Antibacterial Activities ofPolycations with Pendant Biguanide Groups”; Antimicrob. AgentsChemother. 26(2), p139-44 (1984)).

The unexpected nature of the observation that nonleachable binding iscaused by employment of a drying step is further evidenced by the factthat while it is generally known that a strong polyelectrolyte complex(PEC) can be formed between oppositely charged polymers, formation ofsuch PECs generally proceeds spontaneously, even in aqueous solution.For instance, aqueous solutions of oppositely charged polyelectrolytesmay be mixed, resulting in immediate precipitation of a solid PEC. Forinstance, in U.S. Pat. No. 6,776,985, Saller reports the formation ofPEC microcapsules formed by reaction of aqueous solutions of cellulosesulphate and polyDADMAC. These solid PEC materials are formed withinmilliseconds upon mixing of the two aqueous solutions, without the needfor drying or removal of water. It should be noted that cellulosesulphate is substantially different substance than the substratematerials of the current invention (such as cotton), in that cellulosesulphate is a highly ionic substance which is soluble in water.

The inherently antimicrobial materials produced using the currentinventive method are distinct from those described by Senka (JP09078785, see above discussion), in that the inherently antimicrobialmaterials produced by the current invention are produced due to specificinteractions on the molecular level between the substrate and theapplied polymeric antimicrobial cationic polyelectrolyte, as describedabove. In contrast, the materials described by Senka depend on theinteraction of individual copolymer molecules with each other to form asolid insoluble network. In other words, the copolymers described bySenka are capable of undergoing self-crosslinking. The term“self-crosslinking” means that the polymer has the capability to undergoa chemical or physical reaction with itself which results in bridging,bonding, or attachment between its individual polymer chain molecules toform a three dimensional network structure consisting essentially of asingle large molecule, without the need to react with any outsidereagents such as catalysts or crosslinking agents which are not alreadypart of the polymer's molecular structure. The polymer molecules areindividually bonded to the substrate, and hence behave as part of thesubstrate. Once crosslinked, the polymer becomes insoluble; although,the crosslinked polymeric network may be capable of absorbing water toform a gel. The antimicrobial cationic polyelectrolytes of the currentinvention are not self-crosslinking. Since there is no crosslinkedpolymer network to become attached to the surface of the inherentlyantimicrobial materials of the current invention, the materials retainthe general physical properties of the untreated substrates such asappearance, feel, texture, hand, softness, and flexibility. Crosslinkedcoating materials could also tend to cement individual fibers together,or fill-in voids and pores, both of which could adversely affectsubstrate properties. In contrast the materials prepared using acoating, such as described by Senka, may be expected to become stiff,brittle, or non-adherent due to intermolecular association orcrosslinking of the self-crosslinking polymeric antimicrobials used. Asan analogy of the differences between the two types of systems, considerthe differences between a varnish used to finish wood by providing aprotective glossy coating, versus a wood-stain type of product whichmerely colors the wood itself without changing its texture. Varnishmaterials are similar to the materials of Senka, in that they aregenerally self-crosslinking, and once they dry they become insoluble.The dried varnish; however, may be easily peeled away from theunderlying wood surface. The wood-stain, on the other hand, willgenerally be more substantive to the wood substrate and more difficultto remove. The interactions of the wood-stain are only between theindividual stain molecules and the wood substrate. The stain moleculesare more similar to the antimicrobial cationic polyelectrolytes of thecurrent invention in that they do not interact with themselves to form athree dimensional coating.

It should be emphasized that in most processes intended to produce afinal material that is free from leachable additives, it is generallynot intuitive to dry the product before rinsing out any “extra”non-bonded additive. Usually, it will be more cost effective to rinse orwash the material prior to final drying in order to avoid the need fordrying more than once. Many treatment methods (antimicrobial andotherwise) rely on a reaction between the substrate and the appliedadditive, and this reaction usually takes place in solution. Examples ofthis are seen in many of the references cited above (see for instance:Payne, Onabe, Lee, Abel, and Batich). This rinsing step prior to dryingof the treated material is also widely used in textile treatmentprocesses such as dye treatment using disperse, reactive, or vat dyes.In the case of the current invention; however, the reaction between thesubstrate and antimicrobial additive is actually caused to occur by thedrying step. Hence, rinsing or washing cannot be done until after thematerial has been dried, or else the additive would be completelyremoved.

Antimicrobial efficacy may be measured by appropriate methods which willbe familiar to one skilled in the art. In particular, a modified versionof the American Association of Textile Chemists and Colorists (AATCC)Test Method 100 (“Antibacterial Finishes on Textiles: Assessment of”), atest designed to test antibacterial finishes of textile materials isuseful, and is described in the following examples. One skilled in theart will recognize that a significant reduction in the number of viablebacteria should be observed when the antimicrobial material is testedaccording to this method, which utilizes a non-antimicrobial (untreated)material with similar physical properties as a “negative control”.Preferably, the reduction in bacterial levels (vs. negative control)should be a factor of 1000 (a “3-log kill”, or 99.9% reduction). Morepreferably, the reduction in bacterial levels (vs. negative control)should be a factor of 10,000 (a “4-log kill”, or a 99.99% reduction).Even more preferably, the reduction in bacterial levels (vs. negativecontrol) should be a factor of 100,000 (a “5-log kill”, or a 99.999%reduction). Most preferably, the reduction in bacterial levels (vs.negative control) should be a factor of 1,000,000 (a “6-log kill”, or99.9999% reduction). It should be noted that limitations of the testmethod may result in lower numerical reductions of bacterial levels ifthe number of viable bacteria in the negative control is low. Forinstance, if the negative control contains only 500 viable bacteria(colony forming units), a reduction factor of 500 (a 2.7-log kill) isthe maximum possible result; however, in this case the result representsa 100% reduction of bacterial population, and is perfectly acceptable.Generally, when the standard method is followed using absorbent textiletest articles, the growth of most commonly-encountered bacteria in thenegative control will be in the range of 100,000 to 10,000,000 colonyforming units.

Quaternary ammonium compounds, including the non-leachably boundantimicrobials of this invention, generally show greater activityagainst Gram+ bacterial organisms than against Gram− bacterialorganisms. Thus, a higher level of bonded antimicrobial is needed inorder to achieve the same antimicrobial efficacy against Gram−organisms. Surprisingly, it is found that the necessary levels ofnon-leachably bound antimicrobial in materials prepared by the currentinvention are substantially lower than those required when other methodsare used to attach similar antimicrobials to similar substrates. Forinstance, in a specific embodiment of this invention, a cotton substratetreated with a 2% aqueous solution of polyDADMAC, followed by drying andsequential rinsing in saline and deionized water was found to havegreater than 6-log efficacy against a wide range of both Gram+ and Gram−organisms (see examples below for experimental details). Elementalanalysis for nitrogen content revealed that the overall content ofquaternary ammonium antimicrobial cationic polyelectrolyte (polyDADMAC)was less than 1 weight percent (<1 wt %). In contrast, materialsproduced using antimicrobial treatments involving graft copolymerizationof polyDADMAC monomer onto cotton substrates using the methods describedby Batich, U.S. Pat. No. 7,045,673 and application US 20020177828 A1,must contain greater than approximately 5 wt % antimicrobial forequivalent antimicrobial efficacy when the same substrate (cotton), andthe same cationic polyelectrolyte antimicrobial cationic polyelectrolyte( polyDADMAC) is used. A comparative example showing the antimicrobialefficacy of inherently antimicrobial materials made utilizing thegrafting method of Batich is provided below.

Preferably, the antimicrobial composition of the current invention iseffective against Gram+ bacteria. More preferably, the antimicrobial ofthe current composition is effective against Gram+ bacteria, and Gram−bacteria. Most preferably, the antimicrobial of the current compositionis effective against Gram+ bacteria, Gram− bacteria, and also fungal andviral organisms.

When a quaternary ammonium compound is utilized as the antimicrobialagent in the practice of this invention, it is possible to estimate therelative degree of binding of quaternary antimicrobial to the substrateby utilizing a dye test which is described in the examples given below.It is found that there is a positive correlation between the results ofthe dye test described, and the antimicrobial performance. Generally,materials which exhibit a dark blue color result using the specifiedtest procedure, will exhibit excellent antimicrobial activity (>6-logreduction of pseudomonas, for instance), while lighter blue intensityresults correlate to lower antimicrobial efficacy.

In light of the general disclosure provided herein above, with respectto the manner of practicing this inventive method, those skilled in theart will appreciate that this disclosure enables the practice of theinventive method as defined in the attached claims. However, thefollowing experimental details are provided to ensure a complete writtendescription of this invention, including the best mode thereof. However,it will be appreciated that the scope of this invention should not beconstrued in terms of the specific examples provided. Rather, the scopeof this invention is to be apprehended with reference to the claimsappended hereto, in light of the complete description of this inventivemethod constituted by this entire disclosure.

EXAMPLE 1A Treatment of a Rayon Substrate with PolymerizedDiallyldimethylammonium chloride—Leachable Portion Rinsed.

This example demonstrates the binding of a polymeric quaternary compoundto a substrate. A nonwoven rayon gauze substrate was submerged in a 5%aqueous solution of polymerized diallyldimethylammonium chloride. Afterthe substrate was saturated with the polymer solution, the substrate waspressed free of excess solution by rolling. The wetted and pressed rayonsubstrate was then placed into 50° C. drying oven for four hours, untilthe treated rayon substrate was dried thoroughly. To remove any polymerthat did not become non-leachably attached to the treated rayonsubstrate during the drying step, the treated rayon substrate was rinsedrepeatedly with water until conductivity readings of the rinsate equaledthat of the input rinse water, indicating that the rinsate was free ofunbound polymer. To verify the attachment of cationic polymer, the zetapotential and dye binding behavior were measured, as described inExample 5, below.

EXAMPLE 1B Treatment of a Rayon Substrate with PolymerizedDiallyldimethylammonium Chloride (Comparative Example to DemonstrateEffect of Drying Step Being Omitted).

The method of Example 1A was followed, except that the drying step wasomitted. This results in no significant attachment of the antimicrobialto the substrate.

EXAMPLE 2 Treatment of a Rayon Substrate with PolymerizedDiallyldimethylammonium Chloride—Leachable Portion Retained.

This example demonstrates the binding of a polymeric quatemary compoundto a substrate. A rayon substrate was submerged in a 5% aqueous solutionof polymerized diallyldimethylammonium chloride. After the substrate wassaturated with the polymer solution, the substrate was pressed free ofexcess solution by rolling. The wetted and pressed rayon substrate wasthen placed into 50° C. drying oven for four hours, until the treatedrayon substrate was dried thoroughly.

EXAMPLE 3A Treatment of a Cotton Material Suitable for TextileApplications with Polymerized DiallyldimethylammoniumChloride—Laboratory Scale Production.

This example demonstrates the binding of a polymeric quaternary compoundto a substrate. A knitted cotton fabric substrate was submerged in a 5%aqueous solution of polymerized diallyldimethylammonium chloride. Afterthe substrate was saturated with the polymer solution, the substrate waspressed free of excess solution by rolling. The wetted and pressedcotton substrate was then placed into 50° C. drying oven for four hours,until the treated cotton substrate was dried thoroughly.

EXAMPLE 3B Treatment of a Cotton Material Suitable for TextileApplications with Polymerized DiallyldimethylammoniumChloride—Laboratory Scale Production—Soluble Portion Removed.

The method of Example 3A was followed. After thorough drying of thetreated cotton substrate, it was washed twice in a large excess of 1%aqueous NaCl solution to remove any polymer that did not becomenon-leachably attached to the treated rayon substrate during the dryingstep. It was then rinsed repeatedly with water until conductivityreadings of the rinsate equaled that of the input rinse water,indicating that the rinsate was free of unbound polymer.

EXAMPLE 3C Treatment of a Cotton Material Suitable for TextileApplications with Polymerized DiallyldimethylammoniumChloride—Laboratory Scale Production—Soluble Portion Removed(Comparative Example Demonstrate the Effect of Drying Step BeingOmitted).

The method of Example 3B was followed, except that the drying step wasomitted. This results in no significant attachment of the antimicrobialto the substrate.

EXAMPLE 4 Treatment of a Cotton Material Suitable for TextileApplications with Polymerized Diallyldimethylammonium Chloride—PlantScale Production.

Treated knitted cotton fabric substrate was prepared in an industrialsetting using equipment typically found in the dye house of a textilemill where dyed or bleached cotton fabric emerges from the dye tumblerwet and is then transferred to a pad machine. In this example, the wetfabric direct from the dye tumbler was passed through the pad machine.The wet fabric then entered the pad bath by passing through extractionrollers, which were set to 350 psi and caused the fabric to lose some,if any, of the water it contained. Within the pad bath the fabric wasimmersed in a 5% aqueous solution of polymerized diallyldimethylammoniumchloride. The treated fabric then exited the pad bath and passed throughnip rollers, which were set to 150 psi. The treated fabric was thentransited to a three-stage propane dryer with each stage set to 280° F.and with a transit time for the fabric of 60 seconds, which dried thetreated fabric thoroughly. To remove any polymer that did not becomenon-leachably attached, the treated fabric substrate was rinsed using adye tumbler with repeated fill and rinse cycles with water heated to 60°C., until the measured conductivity of the rinsate equaled that of theinput rinse water.

EXAMPLE 5 Verification of Attachment of an Antimicrobial Polyelectrolyteto the Substrate using a BTB Dye Assay.

The pH indicator dye bromothymol blue (BTB) was used to test for thesuccessful attachment of an antimicrobial polyelectrolyte to asubstrate. This dye assay is best suited for use on substrates that havea neutral or negative zeta potential prior to treatment, such as cotton,rayon, and alginates because binding of the dye to the negativelycharged (untreated) substrates is essentially zero; whereas thenegatively charged dye molecule binds strongly to positively chargedsurfaces, such as those containing bonded non-leachable antimicrobialquaternary ammonium compounds. This method was used to test samplestypical of those produced using the methods described in Examples 1A,1B, 3B and 3C. Each sample was placed into a separate beaker andsaturated with 0.5 wt % BTB dye solution that had been adjusted to pH>10with ammonia. The samples were then rinsed repeatedly with water, untilthe rinsate no longer visibly contained any BTB dye. After the finalrinse, the materials produced in Examples 1A and 3B (with drying step)were substantially blue in appearance, indicating the successfulnon-leachable attachment of the antimicrobial polyelectrolyte, while thesamples produced in Examples 1B and 3C (no drying step) did not show anyblue coloration, indicating the lack of any bonded antimicrobialquaternary material. It should be noted that the blue color of thematerial of Example 3B was significantly darker than that of Example 1A.This is an indication that a higher degree of bonding of antimicrobialquaternary compound is achieved when a cotton substrate is used asopposed to a rayon substrate, when the antimicrobial cationicpolyelectrolyte employed is polymerized diallyldimethylammoniumchloride. Note that depending on the pH of the rinse water, the dyecolor may appear either blue or greenish-yellowish. Adding a few dropsof ammonia to the rinse water will convert the dye color to the blueform. In the practice of this invention, it is useful to evaluate therelative intensity of the final blue color (if any) according to thefollowing scale: Not Blue, Very Light Blue, Light Blue, Blue, Dark Blue,Very Dark Blue. Although this is only a qualitative scale, the resultscan be quantified to some extent by comparison to standard samples, orby using standard methods of measuring color intensity that would beknown to one skilled in the art. In general, it is found that theantimicrobial efficacy correlates to the intensity of the blue color,with samples showing only Very Light or Light Blue color having only lowantimicrobial efficacy.

EXAMPLE 6 Verification of Attachment of the Cationic Compound to theSubstrate Using a Zeta Potential Measurement, and Correlation of ZetaPotential and Antimicrobial Efficacy to Concentration of AntimicrobialPolymer Used in Treatment Solution.

Zeta potential is a measure of the surface charge of a material. Thezeta potential of a successfully treated substrate should besignificantly higher than that of an untreated substrate, due to thepresence of the antimicrobial cationic polyelectrolyte. The zetapotential of each sample was determined by measuring the streamingpotential of the fabric using a Brookhaven-Paar Physica EKAElectrokinetic Analyzer. Each sample was loaded individually into thecylindrical cell attachment forming a fiber plug in the cell. Thedistance between the Ag/AgCl electrodes was adjusted to 30 mm. Bufferedmillimolar potassium chloride was used as the streaming fluid. Eightmeasurements of the streaming potential were made for each sample. Thereported zeta potential is an average of those calculated using eachmeasured streaming potential value. To determine whether surfaceconduction had an effect on the measured streaming potential, thestreaming fluid was then replaced with buffered 0.1M KCl and the zetapotential with surface conduction correction was determined. Anuntreated control sample was included for comparison. Approximately 2.5to 3.0 g of treated or untreated rayon substrate material was compactedinto the measurement cell and the electrodes were set to a specifiedseparation. The fluid utilized by the instrument is for surfaceconduction correction measurements. These solutions were used both assupplied and with 7.4 pH buffer added to determine the streamingpotential at physiologic fluid pH and to minimize pH drift. The surfaceconduction measurements were made to determine the effect of surfaceconduction on the measured zeta potential. For substrates treated athigh polymer concentrations by the method of the current invention, thezeta potential measurements with surface conduction correction aresignificantly greater than without the correction, but the differencebetween these measurements decreases with polymer concentration onsubstrate. The data collected are tabulated as streaming potentials.Untreated cotton substrates have shown a typical zeta potential of −15to −20 mV. Cotton substrates treated by the methods of this inventionhave been demonstrated to show zeta potential values above +15 mV. Rayonproducts show similar values. The corrected zeta potential of anuntreated sample of rayon substrate under pH 7.4 buffered conditions wasin the range of −10 mV, while the treated sample yielded values between+10 and +30 mV.

Samples of 100% knitted jersey material were prepared according to themethod of Examples 3B. The concentration of polyDADMAC in the treatmentsolution was varied from zero to 1.90 wt %. Zeta potentials weremeasured as described above. The antimicrobial efficacy of the materialsagainst Staph. aureus and E. coli was determined by the method describedin Example 7. The results are shown below in Table 1.

TABLE 1 Zeta Potential and Antimicrobial Efficacy of Treated Samples %pDADMAC in Zeta potential Zeta potential Log Reduction Log Reductiontreatment solution (mV) with SCC (mV) of S. aureus of E. coli 1.90% 19.9± 0.8 27.9 ± 0.5 6.19 6.76 0.98% 29.4 ± 0.3 33.5 ± 0.4 6.19 6.76 0.66%24.5 ± 1.9 28.2 ± 2.2 6.19 6.43 0.49% 28.9 ± 0.2 35.2 ± 0.2 6.19 4.840.40% 25.5 ± 0.3 29.1 ± 0.6 5.19 5.99 0.33% 16.2 ± 0.4 18.5 ± 0.5 2.991.80 0.28% 17.0 ± 0.2 19.7 ± 0.3 5.09 1.73 0.25% 16.3 ± 0.1 19.2 ± 0.21.87 1.91 0.22% 11.4 ± 0.2 13.8 ± 0.3 2.40 1.57 0.20% 10.7 ± 0.3 12.5 ±0.3 2.45 1.22 0.00% 24.3 ± 1.7 34.1 ± 0.2 0.00 0.00

EXAMPLE 7 Microbiological Assay to Verify the Antimicrobial Property ofTreated Substrate Material.

Antimicrobial activity of materials prepared using the various methodsand embodiments of this invention were assayed using a modified versionof the American Association of Textile Chemists and Colorists (AATCC)Test Method 100 (“Antibacterial Finishes on Textiles: Assessment of”), atest designed to test antibacterial finishes of textile materials.Overnight cultures (ONC) of test microorganisms were generated inappropriate culture medium using standard methods. Using the ONC, aninoculum solution was prepared containing the test microorganism dilutedto ˜10⁶ CFU/ml in phosphate buffered saline (PBS) and fetal bovine serum(FBS), at 10% v/v. Treated substrate materials (samples) and untreatedsubstrate control materials (controls) were cut into 2.5 cm squares andautoclaved at 121° C. for 30 minutes to eliminate pre-existing microbialcontamination. After autoclaving, samples and controls were allowed tocool for 15 minutes at room temperature. Samples and controls were eachinoculated with 500 μL of inoculum and incubated at 37° C. in sterilecovered petri dishes. After 24 hours incubation, the samples andcontrols were harvested with sterile forceps, placed into separate 15 mLtubes containing 15 mL PBS, and vortexed for 30 seconds to suspend anyremaining viable microorganisms into solution. Appropriate tenfolddilutions of these suspensions were made using PBS solution and spreadonto bacteria culture plates containing growth medium appropriate forthe desired organisms and then incubated overnight at 37° C. Afterovernight culture, colonies growing on each plate are enumerated todetermine antimicrobial efficacy. Data are reported as % killed or logreduction as compared to untreated controls. The dilution, spreading,plating and enumeration were conducted using standard microbiologicaltechniques. Results of this testing on samples made using variousembodiments of the described invention are presented in Table 3.

EXAMPLE 8 Microbiological Assay to Verify the Antimicrobial Property ofa Treated Substrate was Non-Leachably Attached.

Materials prepared using the method of Example 3B, as well as untreatedcontrol materials were rinsed repeatedly to remove unbound antimicrobialcationic polyelectrolyte by the procedure described in Example 3B.Control and sample were cut into 120 sq-cm pieces. Four sample and fourcontrol pieces were placed into separate beakers containing 20 ml of0.9% NaCl solution. Beakers were then placed in an oven at each of thefollowing four conditions: 121° C. for 1 hour, 70° C. for 24 hours, 50°C. for 72 hours, or 37° C. for 120 hours. After the indicated amount oftime, the beakers were removed from the oven and the extract solutionfrom each beaker was harvested. The extract solutions were diluted to ¼log, ½ log, ¾ log and 1 log. Bacteria culture plates were prepared bylawn-spreading a 1/100 dilution of an ONC of test microorganisms, bothS. aureus and E. coli. After the culture medium had absorbed theinoculum, a 20 μl aliquot of each dilution of each extract solution wasplaced at marked locations on the plates. Bacterial culture plates werethen incubated overnight at 37° C. and inspected for observed % kill andsize of the zone of inhibition surrounding marked spots. No evidence ofantimicrobial effect was observed at the marked locations, indicatingthe nonleachable character of the bound microbicide.

EXAMPLE 9 Treatment of a Nonwoven Rayon Fabric Substrate withNonpolymeric Quaternary Ammonium Compounds (Comparative Example).

This example demonstrates the lack of non-leachable binding of anon-polymeric quaternary compound to a cotton substrate and/or lack ofbinding of antimicrobial non-polymeric quaternary compounds to asubstrate. A woven 100% cotton gauze material was used (Kerlix™,manufactured by Kendall) was treated with the non-polymeric quaternaryammonium compounds listed below according to the following procedure.After the substrate was saturated with the solution (5% quaternarycompound), the substrate was pressed free of excess solution. The wettedand pressed substrate was then placed into 80° C. drying oven for 2hours, until the treated substrate was dried thoroughly. After thoroughdrying of the treated cotton substrate, it was washed twice in a largeexcess of 1% aqueous NaCl solution to remove any quaternary compoundthat did not become non-leachably attached to the substrate during thedrying step. It was then rinsed repeatedly with water until conductivityreadings of the rinsate equaled that of the input rinse water,indicating that the rinsate was free of quaternary compound. Thefollowing non-polymeric quaternary ammonium compounds were used:

Sample # compound 9A chlorhexidine 9B tetraethylammonium bromide 9Ctetramethylammonium bromide 9D benzalkonium chloride 9Etetrabutylammonium bromide 9F didecyldimethylammonium chloride

These treated materials were tested for antimicrobial efficacy accordingto the procedure given in Example 7 using Pseudomonas aeruginosa as thetest organism. The antimicrobial efficacy for each sample is listedbelow:

Sample # log reduction 9A >6 9B 0 9C 0 9D 2 9E 5

Fresh portions of the three samples which showed antimicrobial activity(9A, 9D, and 9E) were subjected to extraction testing in order todetermine leachability of the non-polymeric antimicrobials. The methodof Example 8 was used, with the following modifications: Four grams ofmaterial was extracted into 20 mL of 1% NaCl (aq), for 24 hours at 70°C. Undiluted extracts were plated. Activity was observed for all threesamples, indicating lack of non-leachable attachment of non-polymericantimicrobial cationic polyelectrolyte. Note that this is in contrast tothe behavior observed in Example 8 for quaternary antimicrobial cationicpolyelectrolyte.

EXAMPLE 11 Treatment of Laboratory Filter Paper with PolymerizedDiallyldimethylammonium Chloride.

A sample of laboratory filter paper (Whatman) was soaked in an aqueoussolution of 1 wt % polymerized diallyldimethylammonium chloride. Afterthe filter paper substrate was saturated, it was placed into an 80° C.drying oven for two hours, until the treated rayon substrate was driedthoroughly.

After drying, the treated filter paper sample was rinsed three times indeionized water and dried again at 80° C. in the manner described above.A portion of this treated substrate was retained and hereafter isreferred to as Sample 1. Another portion was rinsed two times with 1%sodium chloride solution, followed by three additional rinses withdeionized water, then dried again at 80° C. for two hours. A portion ofthis treated substrate was retained and hereafter is referred to asSample 2.

Portions of Samples 1 and 2 were soaked in a solution 0.5% BTB indicatoradjusted to pH>20 and rinsed repeatedly with deionized water. A uniformmedium blue color, indicative of quaternary ammonium groups bonded tothe paper surface, was imparted to the paper and was not diminished byfurther rinsing or soaking in water. An untreated filter paper controlwas also subjected to the dye test. The untreated filter paper remainedslightly blue at first, however, the blue color leached out aftersoaking in distilled water overnight, and the paper became white.

EXAMPLE 12 Treatment of Cornstarch with PolymerizedDiallyldimethylammonium Chloride.

Fifty grams of cornstarch (Argo) was mixed with 50 mL of a 1 wt %aqueous solution of polymerized diallyldimethylammonium chloride. Themixture was then spread on a pan and allowed to dry at room temperatureovernight and then was dried thoroughly in an oven at 80° C. for twohours. The treated cornstarch was then ground to a powder consistencyand rinsed several times with distilled water. Centrifugation was usedto assist in the recovery of the treated cornstarch powder betweenrinsings. After rinsing, the powder was allowed to dry at roomtemperature overnight and then was dried thoroughly in an oven at 80° C.for two hours. The treated and rinsed cornstarch was then ground to apowder consistency.

A BTB dye test was performed using the method described in Example 5.The treated cornstarch showed a distinct blue color, compared withuntreated starch, which was only a very pale blue.

The antimicrobial activity of both rinsed and un-rinsed cornstarchpowder was assayed by the ability of bacteria cultures to grow insuspension in the presence treated and untreated cornstarch. Treatedsample and untreated control cornstarch material (0.5 g) was placed intoof solution of 9 ml of phosphate buffered saline and 1 ml of a ten-folddilution of ONC of bacteria. Sample and control were shaken on shaker at37° C. for the indicated amount of time. Sample and control were thenserially diluted and the dilutions were streaked onto bacterial cultureplates. After overnight culture at 37° C., colonies growing on eachplate were enumerated. The results are shown in Table 2 below:

TABLE 2 Untreated Time shaken Rinsed samples Unrinsed samples controls(min) Log Reduction CFU/mL 10 3.76 6.91* 8.10E+06 30 4.18 6.65* 4.50E+0660 4.78 6.48* 3.00E+06 120 5.04 6.74* 5.50E+06 *= Complete kill (100%)

EXAMPLE 13 The Following Example Demonstrates the Preparation andProperties of Materials Containing Non-Leachable Bound AntimicrobialsPrepared using Various Polymeric Antimicrobials.

The method of Example 3B was used; however, the following quaternaryammonium polymers were used: polyDADMAC; Poly(vinylbenzyl ammoniumchloride)—otherwise known as pVBTAC;poly(2-(methacryloyloxy)ethyl)trimethylammonium chloride)—otherwiseknown as pTMMC; poly(dimethylaminoethyl methacrylatehydrochloride)—otherwise known as p(DMAEMA). Each polymer wassynthesized from the corresponding monomer in aqueous solution usingfree radical initiators. All polymers were applied to the substrates as5 wt % aqueous solutions. Antimicrobial efficacy was determined by themethod described in Example 7. Results are presented in the followingtable.

Sample log reduction of Pseudomonas aeruginosa pDADMAC 6 pVBTAC 5 pTMMC1 pDMAEMA 1

EXAMPLE 14 Assessment of Effect of Drying Step on Non-LeachableAttachment of Polymeric Antimicrobial to a Cotton Substrate.

A woven 100% cotton gauze material was used as a substrate (Kerlix™,manufactured by Kendall). This substrate was treated with a 5% solutionof polyDADMAC. Drying was conducted in a sealed plastic chamber equippedwith a humidity measuring apparatus (Fisher Scientific model #11-661-18). Samples were left in the chamber at specified humidity (%relative humidity) for a period of approximately 24 hours. Humidity wascontrolled by placing an appropriate amount of wet paper towels on thefloor of the plastic chamber. Samples were suspended on a screen in thecenter of the chamber, approximately 1.5″ above the floor. Samples were“dried” at room temperature. After removal from the humidity chamber,samples were immediately washed according to the procedure of Example3B. Samples were then tested for antimicrobial efficacy according to theprocedure of Example 7. Results are presented below:

Sample log reduction of Pseudomonas aeruginosa 30% rh 8 45% rh 8 70% rh6 90% rh 6 100% rh  2

The degree of dryness of the samples would be expected to be dependenton the relative humidity of the air within the drying chamber. Thus, thepositive effect of thorough drying on non-leachable bonding of polymericquaternary antimicrobial is demonstrated.

EXAMPLE 15 Preparation of Materials with Non-Leachable BoundAntimicrobials.

Materials were prepared according to the method of Example 3B, exceptthat various substrates and polyDADMAC concentrations were used (asindicated below). The products with non-leachably attached antimicrobialpolymer were tested for antimicrobial activity as described in Example7. The organisms tested, and the antimicrobial activity in terms oflog-reduction of viable organisms is presented below:

Full kill Assay # Substrate Organism Treatment Avg. LR (y/n) 275 KerlixCotton Gauze SA 10% PD 7.00 Y 336 Knitted Cotton Jersey SA 0.30% PD 7.34Y 321 Rayon Wound Pad SA 5% PD 6.75 Y 321 Cotton Wound Pad SA 5% PD 6.91Y 272 55% Cotton/45% PET SA 1.0% PD 6.81 Y 217 Bulk cotton SA 2.0% PD7.03 Y 217 Wood pulp SA 2.5% PD 7.03 Y 210 Cotton Socks SA 1.0% PD 6.96Y 169 Bulk Rayon Staple SA 1.25% PD 5.37 N 208 Kerlix Cotton Gauze SE10% PD 7.44 Y 84 Knitted Cotton Jersey SE 0.74% PD 6.81 Y 275 KerlixCotton Gauze EC 10% PD 7.00 Y 258 Knitted Cotton Jersey EC 1.2% PD 7.11Y 110 Cotton Socks EC 1.1% PD 7.53 Y 196 Cotton Gauze EC 4.0% 6.50 Y 169Bulk Rayon Staple EC 1.25% PD 4.73 N 275 Kerlix Cotton Gauze PA 10% PD7.00 Y 330 Jersey Material PA >1% PD 7.0 Y 321 Cotton Wound Pad PA 5% PD7.57 Y 272 55% Cotton/45% PET PA 1.0% PD 5.32 N 225 Wood pulp PA 5.0% PD4.79 N 217 Bulk cotton PA 2.0% PD 7.21 Y 227 Kerlix Cotton Gauze SP1 5%PD 5.81 Y 65 Bulk cotton SP1 5% PD 5.84 N 227 Kerlix Cotton Gauze PV 5%PD 7.53 Y 65 Bulk cotton PV 5% PD 3.65 N 59 Wood pulp PV 1.25% PD 6.56 Y309 Kerlix Cotton Gauze PM 10% PD 6.71 Y 208 Kerlix Cotton Gauze EF 5%PD 7.06 Y 208 Kerlix Cotton Gauze EA 10% PD 5.88 N B 0805 Kerlix CottonGauze MRSA 2% PD 6.06 N 280 55% Cotton/45% PET MRSA ~1.0% PD 6.47 YB0805 Kerlix Cotton Gauze VRE 2% PD 5.20 N B 0805 Kerlix Cotton Gauze LM2% PD 7.54 Y 161 Knitted Cotton Jersey LM ~0.7% PD 6.85 Y 59 Wood pulpLM 1.25% PD 6.65 Y 271 Kerlix Cotton Gauze CX 10% PD 5.68 N 192 CottonSocks CX ~1.6% PD 5.85 Y 177 Knitted Cotton Jersey CX ~0.7% PD 6.00 Y271 Kerlix Cotton Gauze ML 10% PD 6.02 Y 192 Cotton Socks ML ~1.6% PD6.31 Y 174 Knitted Cotton Jersey ML ~0.7% 5.70 Y 169 Bulk Rayon StapleML ~1.25% PD 4.05 Y 84 Knitted Cotton Jersey CD ~0.74% PD 6.26 Y 345Kerlix Cotton Gauze SM 10% PD 6.76 Y 291 Kerlix Cotton Gauze KP 10% PD7.35 Y 65 Bulk cotton KP 5% PD 7.11 Y 51 Knitted Cotton Jersey KP 5% PD6.06 Y 59 Wood pulp SC 1.25% PD 6.67 Y Key: Assay # = internal referencenumber PD = polyDADMAC Avg. LR = Log Reduction Full Kill (y/n) = Yes orNo Organism: SA = Staphylococcus aureus ATCC # 6538 SE = Staphylococcusepidermidis ATCC # 12228 EC = Escherchia coli ATCC # 15597 PA =Pseudomonas aeruginosa ATCC # 15442 SP = Streptococcus pyogenes ATCC #10096 PV = Proteus vulgaris ATCC # 29905 PM = Proteus mirabilis ATCC #7003 EF = Enterococcus faecalis ATCC # 10741 EA = Enterobacter aerogenesATCC # 13048 MRSA = Methicillan resistant S. aureus ATCC # BAA-44 VRE =Vancomycin resistant E. faecium ATCC # 700221 LM = Listeriamonocytogenes ATCC # 13932 CX = Corynebacterium xerosis ATCC # 7711 ML =Micrococcus luteus ATCC # 21102 CD = Corynebacterium diptheriae ATCC #43145 SM = Serratia marcescens ATCC # 13880 KP = Klebsiella pneumoniaeATCC # 13883 SC = Salmonella choleraesuis ATCC # 10708

EXAMPLE 16 A Comparative Example Demonstrating the Preparation andAntimicrobial Efficacy of Materials Prepared by the Method of Batich(U.S. 020177828 A1) (Comparative Example).

A free-radical initiated graft polymerization of DADMAC monomer onto acotton substrate (Kerlix cotton gauze) was carried out according to themethod of Batich et al. Substrates were immersed in an excess of aqueoussolutions containing DADMAC monomer and sodium persulfate initiator (0.9wt %). Solutions were sparged thoroughly with argon gas, then sealed andheated at 60° to 80° C. for at least 4 hours, followed by thoroughrinsing as described in Example 3B to remove unbonded and leachableantimicrobial polymer. The samples were tested for antimicrobialactivity according to the procedure described in Example 7. The testorganism was Pseudomonas aeruginosa. Results are summarized below:

DADMAC conc. Average log reduction 12% −0.28 23% 1.58 35% 4.27 47% 5.0559% 5.72

Comparison with the antimicrobial efficacy of the current invention (asshown in Example 15) demonstrates that the current invention is capableof producing a non-leachable antimicrobial material with greaterefficacy than that provided by the prior art, even when substantiallylower concentrations of antimicrobial agent are used. Furthermore, thiscan be accomplished without the need for conducting the treatment in aninert atmosphere.

EXAMPLE 17 Health & Safety Testing of a Cotton Wound Dressing MaterialContaining a Non-Leachable Antimicrobial Polymer.

Materials prepared according to the method of Example 3B (substrate wasKerlix cotton gauze, and [polyDADMAC] was 5%) were submitted for safetytesting at Toxikon Laboratories, Bedford, Mass. Cytoxicity,Sensitization, Intracutaneous Reactivity, and Acute Systemic ToxicityTests were performed. Such testing is required by regulatory agenciessuch as the FDA before materials such as antimicrobial wound dressingscan be approved for sale and use. The material passed all the tests withlowest (best) possible scores on every test.

Agar diffusion test, ISO 10993-5, 1999. This test is also referred to asa Cytotoxicity test. The material was found to elicit no response fromthe L929 mammalian cell line at 48 hours post exposure to the testarticle: a grade 0 response was found at all time points. The materialwas determined by Toxikon labs to pass this test.

Intracutaneous Injection Test, ISO 10993-10, 2002. The extracts from thematerial were found to elicit no significantly greater biologicalreaction than extracts from negative control articles when tested on NewZealand white rabbits. No signs of toxic response were found at any timepoint, and there was zero incidence of erythema/eschar and zeroincidence of edema, using either for NaCl extract media, or forcottonseed oil extract media. The material was determined by Toxikonlabs to pass this test.

Kligman Maximization Test (also known as sensitization), ISO10993-10993-10, 2002. The material extract elicited no reaction at thechallenge (0% sensitization), following the induction phase. There wereno readings of toxicity, and the scores for all the test animals at alltest time points were 0. As defined by the scoring system of Kligman,this is categorized as a grade I reaction, and the test article isclassified as having weak allergenic potential. The Grade Isensitization is not considered significant according to this test. Thematerial was determined by Toxikon labs to pass this test.

Systemic Injection Test, ISO 10993-11, 1993. The material was found toelicit no significantly greater biological reaction than treated withcontrol articles when tested on Albino Swiss mice. No signs of toxicresponse were found at any time point. The material was determined byToxikon labs to pass this test.

EXAMPLE 18 Determination of Non-Leaching Antimicrobial Polymer Contentof Treated Materials.

Materials were prepared according to the method of Example 3B usingKerlix cotton gauze as the substrate, and varying the concentration ofpolyDADMAC in the treatment solution, as indicated in the table below.Elemental analysis for nitrogen content (Kjelldahl method) was performedby Galbraith Laboratories in Knoxville, Tenn., and is reported as ppm(see table below). Based on the nitrogen content, the % of polyDADMACremaining in the product as a non-leachably attached polymericantimicrobial was calculated by using the formula weight of DADMAC, andsubtracting the nitrogen content of the untreated materials.

% pDADMAC (treatment) Nitrogen (ppm) % PD (product) 0.00% 158 0.00%0.00% 120 0.00% 0.55% 252 0.13% 1.10% 312 0.20% 2.36% 390 0.29% 3.99%520 0.44% 10.00% 598 0.53%

EXAMPLE 19 Demonstration of Retention of Antimicrobial Efficacy ofInherently Antimicrobial Material after Fifty Laundering Cycles.

A sample was prepared according to the procedure of example 3A, exceptthat the treatment solution consisted of 1% polyDADMAC in water. Afterdrying, the product was laundered according to AATCC standard method#135-2110 (Dimensional Changes in Automatic Home Laundering of Woven andKnit Fabrics) to achieve the equivalent of 50 home laundering cycles.The material was then tested for antimicrobial efficacy according to themethod of Example 7, and found to give a 6.6 log reduction of Staphaureus, indicating retention of appreciable inherent antimicrobialactivity, even after fifty laundering cycles. The laundering treatmentwas performed by Textile Testing Service, University of Manitoba(Manitoba, Canada).

EXAMPLE 20 Comparative Example.

Samples of bulk cotton material were treated according to the method ofOnabe (“Studies of Interfacial Properties of Polyelectrolyte-CelluloseSystems. I. Formation and Structure of Adsorbed layers of CatioinicPolyelectrolyte-(Poly-DMDAAC) on Cellulose Fibers” Journal of AppliedPolymer Science. Vol. 22, 3495-3510 (1978). Cotton samples (0.4 gramseach) were immersed in separate 40 mL portions of 2% polyDADMACsolution. Samples were left to soak for either 24 hours, or forapproximately 5 minutes. Samples for each soaking time were thenthoroughly dried (according to the method of the current invention, butdistinct from the method of Onabe). These dried samples, along withduplicate samples subjected to the same soaking conditions describedabove, were then rinsed according to the method of Onabe. Specifically,each sample was placed into a glass filter and rinsed with 1 liter ofdeionized water, then transferred to a glass beaker filled with 200 mLof distilled water, and allowed to stand overnight. The electricalconductivity of the water in each beaker was then measured, with theflowing results:

24 hour/rinsed: 875

24 hour/dried & rinsed: 4

5 minute/rinsed: 8

5 minute/dried & rinsed: 3

The conductivity of the solution is reflective of the concentration ofpolyDADMAC in solution, and indicates two things. First, drying prior torinsing has a pronounced effect, causing a significant reduction in theamount of polyDADMAC leaching (leaving the cotton and migrating intosolution). Second, it appears that the longer soaking time results ingreater absorption of polyDADMAC into the wet cellulose fiber, however,this greater amount is subsequently lost during rinsing, if the rinsingis done prior to drying the sample.

The samples were then thoroughly dried and subjected to the dye testdescribed in Example 5, and then photographed. The results of the dyetesting were that both non-dried rinsed samples showed only a Light Bluecolor, indicative of relatively low, if any, antimicrobial efficacy.Both of the samples subjected to a drying step prior to rinsing showed aDark Blue color, indicative of high antimicrobial efficacy.

1-44. (canceled)
 45. A method of manufacture of an inherentlyantimicrobial material comprising, in sequence, the steps of: a)providing a substrate for use in textiles, medical applications,filters, absorbent materials, or packaging materials, comprised in wholeor in part of a natural or synthetic substrate material. b) exposing thesubstrate to an aqueous solution of an antimicrobial cationicpolyelectrolyte wherein the antimicrobial cationic polyelectrolyte hasan average degree of polymerization of at least 3, and an excesscationic charge density of at least 1 mole per 22,000 grams; wherein thesolution concentration of the antimicrobial cationic polyelectrolyte isat least 0.01 weight percent, and c) thoroughly drying the exposedsubstrate to effect non-leachable bonding between at least a portion ofthe antimicrobial cationic polyelectrolyte and the substrate.
 46. Themethod of claim 45 wherein said substrate is comprised, in whole or inpart, of a natural substrate material selected from the group consistingof polysaccharide, gelatin, chitin, chitosan, protein, collagen,alginate, starch, cotton, wool, silk, and rubber.
 47. The method ofclaim 45 wherein the natural substrate is comprised, in whole or inpart, of a synthetic substrate material selected from the groupconsisting of polyester, polyglycolide, polylactide, rayon, nylon andother polyamides, rubber, acrylic, and polyurethane.
 48. The method ofclaim 45 wherein the natural or synthetic substrate material is gelatin,collagen, starch, cotton, wool, silk, rayon, or rubber.
 49. The methodof claim 45 wherein the natural or synthetic substrate materialcomprises a woven fabric or woven textile.
 50. The method of claim 45wherein the natural or synthetic substrate material comprises anon-woven fabric or non-woven textile.
 51. The method of claim 45wherein said antimicrobial cationic polyelectrolyte comprises polyDADMAChomopolymer or polyVBTAC homopolymer.
 52. The method of claim 51 whereinsaid polyDADMAC homopolymer or polyVBTAC homopolymer has an averagedegree of polymerization of from 100 to 2,500.
 53. The method of claim51 wherein the solution concentration of polyDADMAC homopolymer orpolyVBTAC homopolymer is between 0.05 and 5 weight percent.
 54. Themethod of claim 45 wherein said antimicrobial cationic polyelectrolytehas a minimum excess cationic charge density of at least 1 mole per 212grams of antimicrobial cationic polyelectrolyte.
 55. The method of claim45 wherein the solution concentration of antimicrobial cationicpolyelectrolyte is between 0.05 and 5 weight percent.
 56. The method ofclaim 45 wherein thorough drying is accomplished by application ofinfrared heat, radiant heat, or hot air.
 57. The method of claim 45wherein the substrate comprised in whole or in part of a natural orsynthetic substrate material is gauze, a wound dressing, or a componentof a wound dressing.
 58. The method of claim 45 wherein the inherentlyantimicrobial material is formed into all or part of a wound dressing, aburn dressing, a sanitary pad, a tampon, an intrinsically antimicrobialabsorbent dressing, a diaper, a sanitary wipe, a sponge, a cotton swab,a surgical gown, an isolation gown, a lab coat, a glove, surgicalscrubs, a head cover, a hair cover, a face mask, a suture, a floor mat,a lamp handle cover, a cast liner, a splint liner, padding, gauze,sterile packaging, a mattress cover, bedding, a sheet, a towel,clothing, underwear, a sock, shoe-cover, an automobile air filter, anairplane air filter, an HVAC system air filter, a military protectivegarment, an apparatus for protection against a biohazard or biologicalwarfare agent, apparel for food handling, a surface for foodpreparation, a contact lens, or carpet.
 59. The method of claim 45 whichincludes the additional step of rinsing, washing, or extracting toremove any leachable unbonded portion of said antimicrobial cationicpolyelectrolyte from the inherently antimicrobial material.
 60. A methodof manufacture of an inherently antimicrobial material comprising, insequence, the steps of: a) providing a substrate for use in textiles,medical applications, filters, absorbent materials, or packagingmaterials, comprised in whole or in part of a cellulosic material, b)exposing the substrate comprised in whole or in part of a cellulosicmaterial to an aqueous solution of an antimicrobial cationicpolyelectrolyte wherein the antimicrobial cationic polyelectrolyte hasan average degree of polymerization of at least 3, and an excesscationic charge density of at least 1 mole per 22,000 grams; wherein thesolution concentration of the antimicrobial cationic polyelectrolyte isat least 0.01 weight percent, c) thoroughly drying the exposedsubstrate, and d) rinsing, washing, or extracting to remove anyleachable unbonded portion of said antimicrobial cationicpolyelectrolyte from the inherently antimicrobial material. wherebynon-leachable bonding is effected between at least a portion of theantimicrobial cationic polyelectrolyte and the substrate comprised inwhole or in part of a cellulosic material.
 61. The method of claim 60wherein said antimicrobial cationic polyelectrolyte has an averagedegree of polymerization of from 20 to 10,000.
 62. The method of claim60 wherein said antimicrobial cationic polyelectrolyte has a minimumexcess cationic charge density of at least 1 mole per 162 grams ofantimicrobial cationic polyelectrolyte.
 63. The method of claim 60wherein said antimicrobial cationic polyelectrolyte has a minimum excesscationic charge density of at least 1 mole per 212 grams ofantimicrobial cationic polyelectrolyte.
 64. The method of claim 60wherein said antimicrobial cationic polyelectrolyte is an ammoniumcompound
 65. The method of claim 60 wherein said antimicrobial cationicpolyelectrolyte is a phosphonium compound.
 66. The method of claim 60wherein the solution concentration of antimicrobial cationicpolyelectrolyte is between 0.05 and 5 weight percent.
 67. The method ofclaim 60 wherein thorough drying is accomplished by application ofinfrared heat, radiant heat, or hot air.
 68. A method of manufacture ofan inherently antimicrobial absorbent material comprising, in sequence,the steps of: a) providing an absorbent substrate for use in textiles,medical applications, filters, absorbent materials, or packagingmaterials, comprised in whole or in part of cotton, b) exposing thesubstrate comprised in whole or in part of cotton to an aqueous solutioncomprising polyDADMAC homopolymer or polyVBTAC homopolymer; wherein thepolyDADMAC homopolymer or polyVBTAC homopolymer has an average degree ofpolymerization of at least 3, and an excess cationic charge density ofat least 1 mole per 212 grams; wherein the solution concentration ofpolyDADMAC homopolymer or polyVBTAC homopolymer is at least 0.01 weightpercent, and c) thoroughly drying the exposed substrate, wherebynon-leachable bonding is effected between at least a portion of theantimicrobial cationic polyelectrolyte and the substrate comprised inwhole or in part of cotton.
 69. The method of claim 68 wherein theabsorbent substrate is used in textiles.
 70. The method of claim 68wherein the absorbent substrate is used in medical applications.
 71. Themethod of claim 68 wherein the absorbent substrate is used in absorbentmaterials.
 72. The method of claim 68 wherein the substrate comprises inwhole, or in part gauze, a wound dressing, or a component of a wounddressing.
 73. The method of claim 68 wherein the substrate comprises inwhole, or in part, a fabric or a textile.
 74. The method of claim 68wherein the inherently antimicrobial material is formed into all or partof a wound dressing, a burn dressing, a sanitary pad, a tampon, anintrinsically antimicrobial absorbent dressing, a diaper, a sanitarywipe, a sponge, a cotton swab, a surgical gown, an isolation gown, a labcoat, a glove, surgical scrubs, a head cover, a hair cover, a face mask,a suture, a floor mat, a lamp handle cover, a cast liner, a splintliner, padding, gauze, sterile packaging, a mattress cover, bedding, asheet, a towel, clothing, underwear, a sock, shoe-cover, an automobileair filter, an airplane air filter, an HVAC system air filter, amilitary protective garment, an apparatus for protection against abiohazard or biological warfare agent, apparel for food handling, asurface for food preparation, or carpet.
 75. The method of claim 68which includes the additional step of rinsing the inherentlyantimicrobial material to remove any leachable unbonded portion of saidpolyDADMAC homopolymer or polyVBTAC homopolymer.
 76. The method of claim68 wherein said polyDADMAC homopolymer or polyVBTAC homopolymer has anaverage degree of polymerization of from 100 to 2,500.
 77. The method ofclaim 68 wherein the solution concentration of polyDADMAC homopolymer orpolyVBTAC homopolymer is between 0.05 and 5 weight percent.
 78. Themethod of claim 68 wherein thorough drying is accomplished byapplication of infrared heat, radiant heat, or hot air.
 79. A method ofattaching an antimicrobial cationic polyelectrolyte to a substrate foruse in textiles, medical applications, filters, absorbent materials, orpackaging materials, wherein said substrate is comprised in whole or inpart of hydroxyethyl cellulose, carboxymethyl cellulose, methylcellulose, polysaccharide, gelatin, chitin, chitosan, alginate, starch,collagen, polyglycolide, polylactide, polyamide, polyurethane, rubber,polyester, acrylic, nylon, rayon, silk, linen, cotton, wool, wovenflexible material, fabric, protein, collagen, or absorbent materials foraqueous fluids, and wherein said antimicrobial cationic polyelectrolyteis not self-crosslinking and has an average degree of polymerization ofat least 3, wherein said method comprises the steps of wetting thesubstrate with an aqueous solution of the antimicrobial cationicpolyelectrolyte followed by drying of the wetted substrate, wherein saiddrying causes at least a portion of the antimicrobial cationicpolyelectrolyte to become attached to the substrate in a non-leachablemanner, and wherein the attached antimicrobial cationic polyelectrolyteprovides non-leaching antimicrobial activity to the resulting product.80. The method of claim 79 wherein the substrate comprises in whole, orin part, a fabric or a textile.
 81. The method of claim 79 wherein thesubstrate is formed into all or part of a wound dressing, a burndressing, a sanitary pad, a tampon, an intrinsically antimicrobialabsorbent dressing, a diaper, a sanitary wipe, a sponge, a cotton swab,a surgical gown, an isolation gown, a lab coat, a glove, surgicalscrubs, a head cover, a hair cover, a face mask, a suture, a floor mat,a lamp handle cover, a cast liner, a splint liner, padding, gauze,sterile packaging, a mattress cover, bedding, a sheet, a towel,clothing, underwear, a sock, shoe-cover, an automobile air filter, anairplane air filter, an HVAC system air filter, a military protectivegarment, an apparatus for protection against a biohazard or biologicalwarfare agent, apparel for food handling, a surface for foodpreparation, a contact lens, or carpet.
 82. The method of claim 79further comprising rinsing, washing, or extracting to remove anyleachable unbonded portion of said antimicrobial cationicpolyelectrolyte from the inherently antimicrobial material.
 83. Themethod of claim 79 wherein drying is accomplished by application ofinfrared heat, radiant heat, or hot air.
 84. The method of claim 79wherein the substrate is comprised in whole or in part of cotton, andwherein the antimicrobial cationic polyelectrolyte that is not selfcrosslinking comprises polyDADMAC homopolymer having an excess cationiccharge density of at least 1 mole per 162 grams or polyVBTAC homopolymerhaving an excess cationic charge density of at least 1 mole per 212grams; and wherein the solution concentration of polyDADMAC homopolymeror polyVBTAC homopolymer is at least 0.01 weight percent.
 85. The methodof claim 84 wherein said polyDADMAC homopolymer or polyVBTAC homopolymerhas an average degree of polymerization of from 100 to 2,500.
 86. Themethod of claim 84 wherein the solution concentration of polyDADMAChomopolymer or polyVBTAC homopolymer is between 0.05 and 5 weightpercent.